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Sample records for magnetic bioplex assay

  1. T helper (Th) 1 and Th2 cytokine expression profile in dengue and malaria infection using magnetic bead-based bio-plex assay.

    PubMed

    Maneekan, Pannamas; Leaungwutiwong, Pornsawan; Misse, Dorothee; Luplertlop, Natthanej

    2013-01-01

    Dengue and malaria infections are two very common vector-borne diseases annually affecting millions of people around the world. Both diseases show a variety of clinical presentations, ranging from mild symptoms of dengue fever (DF) to severe dengue hemorrhagic fever (DHF) in dengue infection, and low and high parasitemia in malaria infection. T helper (Th)1 and Th2 cytokine expressions in mild and severe forms of dengue virus type-2 (DENV-2) and Plasmodium falciparum infection, were compared to normal human sera using high throughput magnetic bead-based Bio-Plex assay. A significant elevation of Th1 and Th2 cytokines expression [interleukin (IL)-2, IL-4, IL-5, IL-10, IL-13, granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha] was detected in DENV-2 and P. falciparum malaria infections compared with normal controls (p < 0.05). DENV-2 infection showed a slight higher expression of Th1 and Th2 cytokines in DHF than DF, except for IL-13. In P. falciparum infection, high parasitemia showed a significantly higher expression of IL-4, IL-10, GM-CSF, and TNF-alpha (p < 0.05). Both DENV-2 and P. falciparum malaria infections manifested high IL-10 expression, greatest among the cytokines examined, and in the severe forms of infection. The results of this study should lead to a better understanding of pathogenesis of dengue infection and P. falciparum malaria.

  2. Magnetic bead based immuno-detection of Listeria monocytogenes and Listeria ivanovii from infant formula and leafy green vegetables using the Bio-Plex suspension array system.

    PubMed

    Day, J B; Basavanna, U

    2015-04-01

    Listeriosis, a disease contracted via the consumption of foods contaminated with pathogenic Listeria species, can produce severe symptoms and high mortality in susceptible people and animals. The development of molecular methods and immuno-based techniques for detection of pathogenic Listeria in foods has been challenging due to the presence of assay inhibiting food components. In this study, we utilize a macrophage cell culture system for the isolation and enrichment of Listeria monocytogenes and Listeria ivanovii from infant formula and leafy green vegetables for subsequent identification using the Luminex xMAP technique. Macrophage monolayers were exposed to infant formula, lettuce and celery contaminated with L. monocytogenes or L. ivanovii. Magnetic microspheres conjugated to Listeria specific antibody were used to capture Listeria from infected macrophages and then analyzed using the Bio-Plex 200 analyzer. As few as 10 CFU/mL or g of L. monocytogenes was detected in all foods tested. The detection limit for L. ivanovii was 10 CFU/mL in infant formula and 100 CFU/g in leafy greens. Microsphere bound Listeria obtained from infected macrophage lysates could also be isolated on selective media for subsequent confirmatory identification. This method presumptively identifies L. monocytogenes and L. ivanovii from infant formula, lettuce and celery in less than 28 h with confirmatory identifications completed in less than 48 h. PMID:25475329

  3. Magnetic bead based immuno-detection of Listeria monocytogenes and Listeria ivanovii from infant formula and leafy green vegetables using the Bio-Plex suspension array system.

    PubMed

    Day, J B; Basavanna, U

    2015-04-01

    Listeriosis, a disease contracted via the consumption of foods contaminated with pathogenic Listeria species, can produce severe symptoms and high mortality in susceptible people and animals. The development of molecular methods and immuno-based techniques for detection of pathogenic Listeria in foods has been challenging due to the presence of assay inhibiting food components. In this study, we utilize a macrophage cell culture system for the isolation and enrichment of Listeria monocytogenes and Listeria ivanovii from infant formula and leafy green vegetables for subsequent identification using the Luminex xMAP technique. Macrophage monolayers were exposed to infant formula, lettuce and celery contaminated with L. monocytogenes or L. ivanovii. Magnetic microspheres conjugated to Listeria specific antibody were used to capture Listeria from infected macrophages and then analyzed using the Bio-Plex 200 analyzer. As few as 10 CFU/mL or g of L. monocytogenes was detected in all foods tested. The detection limit for L. ivanovii was 10 CFU/mL in infant formula and 100 CFU/g in leafy greens. Microsphere bound Listeria obtained from infected macrophage lysates could also be isolated on selective media for subsequent confirmatory identification. This method presumptively identifies L. monocytogenes and L. ivanovii from infant formula, lettuce and celery in less than 28 h with confirmatory identifications completed in less than 48 h.

  4. Modeling of Solid Waste Processing Options in BIO-Plex

    NASA Technical Reports Server (NTRS)

    Rodriguez, Luis F.; Finn, Cory; Kang, Sukwon; Hogan, John; Luna, Bernadette (Technical Monitor)

    2000-01-01

    BIO-Plex is a ground-based test bed currently under development by NASA for testing technologies and practices that may be utilized in future long-term life support missions. All aspects of such an Advanced Life Support (ALS) System must be considered to confidently construct a reliable system, which will not only allow the crew to survive in harsh environments, but allow the crew time to perform meaningful research. Effective handling of solid wastes is a critical aspect of the system, especially when recovery of resources contained in the waste is required. This is particularly important for ALS Systems configurations that include a Biomass Production Chamber. In these cases, significant amounts of inedible biomass waste may be produced, which can ultimately serve as a repository of necessary resources for sustaining life, notably carbon, water, and plant nutrients. Numerous biological and physicochemical solid waste processing options have been considered. Biological options include composting, aerobic digestion, and anaerobic digestion. Physicochemical options include pyrolysis, SCWO (supercritical water oxidation), various incineration configurations, microwave incineration, magnetically assisted gasification, and low temperature plasma reaction. Modeling of these options is a necessary step to assist in the design process. A previously developed top-level model of BIO-Plex implemented in MATLAB Simulink (r) for the use of systems analysis and design has been adopted for this analysis. Presently, this model only considered incineration for solid waste processing. Present work, reported here, includes the expansion of this model to include a wider array of solid waste processing options selected from the above options, bearing in mind potential, near term solid waste treatment systems. Furthermore, a trade study has also been performed among these solid waste processing technologies in an effort to determine the ideal technology for long-term life support

  5. BIO-Plex Information System Concept

    NASA Technical Reports Server (NTRS)

    Jones, Harry; Boulanger, Richard; Arnold, James O. (Technical Monitor)

    1999-01-01

    This paper describes a suggested design for an integrated information system for the proposed BIO-Plex (Bioregenerative Planetary Life Support Systems Test Complex) at Johnson Space Center (JSC), including distributed control systems, central control, networks, database servers, personal computers and workstations, applications software, and external communications. The system will have an open commercial computing and networking, architecture. The network will provide automatic real-time transfer of information to database server computers which perform data collection and validation. This information system will support integrated, data sharing applications for everything, from system alarms to management summaries. Most existing complex process control systems have information gaps between the different real time subsystems, between these subsystems and central controller, between the central controller and system level planning and analysis application software, and between the system level applications and management overview reporting. An integrated information system is vitally necessary as the basis for the integration of planning, scheduling, modeling, monitoring, and control, which will allow improved monitoring and control based on timely, accurate and complete data. Data describing the system configuration and the real time processes can be collected, checked and reconciled, analyzed and stored in database servers that can be accessed by all applications. The required technology is available. The only opportunity to design a distributed, nonredundant, integrated system is before it is built. Retrofit is extremely difficult and costly.

  6. Dynamic Model of the BIO-Plex Air Revitalization System

    NASA Technical Reports Server (NTRS)

    Finn, Cory; Meyers, Karen; Duffield, Bruce; Luna, Bernadette (Technical Monitor)

    2000-01-01

    The BIO-Plex facility will need to support a variety of life support system designs and operation strategies. These systems will be tested and evaluated in the BIO-Plex facility. An important goal of the life support program is to identify designs that best meet all size and performance constraints for a variety of possible future missions. Integrated human testing is a necessary step in reaching this goal. System modeling and analysis will also play an important role in this endeavor. Currently, simulation studies are being used to estimate air revitalization buffer and storage requirements in order to develop the infrastructure requirements of the BIO-Plex facility. Simulation studies are also being used to verify that the envisioned operation strategy will be able to meet all performance criteria. In this paper, a simulation study is presented for a nominal BIO-Plex scenario with a high-level of crop growth. A general description of the dynamic mass flow model is provided, along with some simulation results. The paper also discusses sizing and operations issues and describes plans for future simulation studies.

  7. Modeling Separate and Combined Atmospheres in BIO-Plex

    NASA Technical Reports Server (NTRS)

    Jones, Harry; Finn, Cory; Kwauk, Xian-Min; Blackwell, Charles; Luna, Bernadette (Technical Monitor)

    2000-01-01

    We modeled BIO-Plex designs with separate or combined atmospheres and then simulated controlling the atmosphere composition. The BIO-Plex is the Bioregenerative Planetary Life Support Systems Test Complex, a large regenerative life support test facility under development at NASA Johnson Space Center. Although plants grow better at above-normal carbon dioxide levels, humans can tolerate even higher carbon dioxide levels. incinerator exhaust has very high levels of carbon dioxide. An elaborate BIO-Plex design would maintain different atmospheres in the crew and plant chambers and isolate the incinerator exhaust in the airlock. This design easily controls the crew and plant carbon dioxide levels but it uses many gas processors, buffers, and controllers. If all the crew's food is grown inside BIO-Plex, all the carbon dioxide required by the plants is supplied by crew respiration and the incineration of plant and food waste. Because the oxygen mass flow must balance in a closed loop, the plants supply all the oxygen required by the crew and the incinerator. Using plants for air revitalization allows using fewer gas processors, buffers, and controllers. In the simplest design, a single combined atmosphere was used for the crew, the plant chamber, and the incinerator. All gas processors, buffers, and controllers were eliminated. The carbon dioxide levels were necessarily similar for the crew and plants. If most of the food is grown, carbon dioxide can be controlled at the desired level by scheduling incineration. An intermediate design uses one atmosphere for the crew and incinerator chambers and a second for the plant chamber. This allows different carbon dioxide levels for the crew and plants. Better control of the atmosphere is obtained by varying the incineration rate. Less gas processing, storage, and control is needed if more food is grown.

  8. Modeling Separate and Combined Atmospheres in BIO-Plex

    NASA Technical Reports Server (NTRS)

    Jones, Harry; Finn, Cory; Kwauk, Xianmin; Blackwell, Charles; Luna, Bernadette (Technical Monitor)

    2000-01-01

    We modeled BIO-Plex designs with separate or combined atmospheres and then simulated controlling the atmosphere composition. The BIO-Plex is the Bioregenerative Planetary Life Support Systems Test Complex, a large regenerative life support test facility under development at NASA Johnson Space Center. Although plants grow better at above-normal carbon dioxide levels, humans can tolerate even higher carbon dioxide levels. Incinerator exhaust has very high levels of carbon dioxide. An elaborate BIO-Plex design would maintain different atmospheres in the crew and plant chambers and isolate the incinerator exhaust in the airlock. This design easily controls the crew and plant carbon dioxide levels but it uses many gas processors, buffers, and controllers. If all the crew's food is grown inside BIO-Plex, all the carbon dioxide required by the plants is supplied by crew respiration and the incineration of plant and food waste. Because the oxygen mass flow must balance in a closed loop, the plants supply all the oxygen required by the crew and the incinerator. Using plants for air revitalization allows using fewer gas processors, buffers, and controllers. In the simplest design, a single combined atmosphere was used for the crew, the plant chamber, and the incinerator. All gas processors, buffers, and controllers were eliminated. The carbon dioxide levels were necessarily similar for the crew and plants. If most of the food is grown, carbon dioxide can be controlled at the desired level by scheduling incineration. An intermediate design uses one atmosphere for the crew and incinerator chambers and a second for the plant chamber. This allows different carbon dioxide levels for the crew and plants. Better control of the atmosphere is obtained by varying the incineration rate. Less gas processing storage and control is needed if more food is grown.

  9. Comparison of non-magnetic and magnetic beads in bead-based assays.

    PubMed

    Hansenová Maňásková, Silvie; van Belkum, Alex; Endtz, Hubert P; Bikker, Floris J; Veerman, Enno C I; van Wamel, Willem J B

    2016-09-01

    Multiplex bead-based flow cytometry is an attractive way for simultaneous, rapid and cost-effective analysis of multiple analytes in a single sample. Previously, we developed various bead-based assays using non-magnetic beads coated with Staphylococcus aureus and Streptococcus pneumoniae antigens for the detection of antibodies. Here, we compared the performance of the assay using non-magnetic beads with one based on the newly developed magnetic beads. We optimized the magnetic beads' coupling procedure and antibody detection assays for S. aureus and S. pneumoniae antigens and we measured IgG in human pooled serum against a series of S. aureus and S. pneumoniae-derived antigens in a singleplex and in a multiplex assay, respectively. For the multiplex assay, the comparison between magnetic and non-magnetic beads showed: i) in the majority of the cases (13 of the 17 tested S. pneumoniae antigens) significantly higher Median Fluorescence Intensity (MFI) values, ii) lower detection limits, iii) lower coefficient of variation (CV: 12% vs. 7% for non-magnetic vs. magnetic beads), so lower inter-assay variation and hence higher reproducibility. Magnetic bead coupling is cost effective, as we used 25% of the normal amount of antigen and only 50% of the beads in comparison to the non-magnetic beads. This optimized magnetic-based assay, which combines ease of use with an improved assay performance, allows detection of antibodies with a low titer that are potentially missed with the non-magnetic-based assay. PMID:27296810

  10. Safety analysis and hazard control during food processing and storage in the BIO-Plex Interconnecting Transfer Tunnel.

    PubMed

    Hentges, D L

    2000-01-01

    The food system, being designed for the BIO-Plex (Bioregenerative Planetary Life Support Systems Test Complex), will be a plant-based diet that requires most of the food to be grown, processed, and prepared in the BIO-Plex. Conversion of crops to edible foods will require extensive food processing within the closed environment of this habitat. Because all consumables in the BIO-Plex will be recycled and reused, food safety is a primary concern. Multifunctional equipment necessary for food processing of the baseline crops (wheat, soybeans, rice, peanuts, dried beans, potatoes, sweet potatoes, lettuce, chard, tomatoes, green onions, carrots, and radishes) was identified. Recommendations for placement of the food processing equipment in the Interconnecting Transfer Tunnel (ITT) of the BIO-Plex were made to facilitate the processing flow diagrams, increase work efficiency, and prevent cross-contamination of pathogens and antinutrients. Sanitation equipment and procedures necessary during food processing in the ITT are described.

  11. Matching Crew Diet and Crop Food Production in BIO-Plex

    NASA Technical Reports Server (NTRS)

    Jones, Harry; Kwauk, Xianmin; Mead, Susan C. (Technical Monitor)

    2000-01-01

    This paper matches the BIO-Plex crop food production to the crew diet requirements. The expected average calorie requirement for BIO-Plex is 2,975 Calories per crewmember per day, for a randomly selected crew with a typical level of physical activity. The range of 2,550 to 3,400 Calories will cover about two-thirds of all crews. The exact calorie requirement will depend on the gender composition, individual weights, exercise, and work effort of the selected crew. The expected average crewmember calorie requirement can be met by 430 grams of carbohydrate, 100 grams of fat, and 90 grams of protein per crewmember per day, for a total of 620 grams. Some fat can replaced by carbohydrate. Each crewmember requires only 2 grams of vitamins and minerals per day. Only unusually restricted diets may lack essential nutrients. The Advanced Life Support (ALS) consensus is that BIO-Plex should grow wheat, potato, and soybean, and maybe sweet potato or peanut, and maybe lettuce and tomato. The BIO-Plex Biomass Production System food production and the external food supply must be matched to the crew diet requirement for calories and nutritional balance. The crop production and external supply specifications can each be varied as long as their sum matches the required diet specification. We have wide flexibility in choosing the crops and resupply. We can easily grow one-half the crew calories in one BIO-Plex Biomass Production Chamber (BPC) if we grow only the most productive crops (wheat, potato, and sweet potato) and it we achieve nominal crop productivity. If we assume higher productivity we can grow a wider variety of crops. If we grow one-half of the crew calories, externally supplied foods can easily provide the other half of the calories and balance the diet. We can not grow 95 percent of the crew calories in two BPCs at nominal productivity while growing a balanced diet. We produce maximum calories by growing wheat, potato, and peanut.

  12. GMR sensors and magnetic nanoparticles for immuno-chromatographic assays

    NASA Astrophysics Data System (ADS)

    Marquina, C.; de Teresa, J. M.; Serrate, D.; Marzo, J.; Cardoso, F. A.; Saurel, D.; Cardoso, S.; Freitas, P. P.; Ibarra, M. R.

    2012-10-01

    Conventional tests based on immunorecognition and on the use of coloured colloidal particles have still some drawbacks that limit their use: they do not provide a quantitative determination of the analyte, and their sensitivity is limited. Our strategy to overcome these disadvantages consists in the use of superparamagnetic core-shell nanoparticles to tag the analyte. The use of these magnetic labels allows us to quantify the amount of analyte present in our sample with a very high sensitivity, detecting their magnetic response by means of the suitable magnetic sensor. Our method is based on measuring the magnetoresistive response of a spin-valve giant magnetoresistive (GMR) sensor placed in proximity to the magnetic nanoparticles present in the lateral flow strip. Here, a brief description of our prototype and of the measurement procedure will be presented, as well as preliminary assays using our biosensor to detect the hCG pregnancy hormone in a solution. A crucial aspect to take into account in order to increase the sensitivity is the proper functionalisation of the nanoparticle shell, in order to achieve an oriented immobilisation of the antibodies to be used in the immunorecognition process. Several strategies to further increase the sensor sensitivity are suggested.

  13. Bioplex technology: novel synthetic gene delivery pharmaceutical based on peptides anchored to nucleic acids.

    PubMed

    Simonson, Oscar E; Svahn, Mathias G; Törnquist, Elisabeth; Lundin, Karin E; Smith, C I E

    2005-01-01

    Non-viral gene delivery is an important approach in order to establish safe in vivo gene therapy in the clinic. Although viral vectors currently exhibit superior gene transfer efficacy, the safety aspect of viral gene delivery is a concern. In order to improve non-viral in vivo gene delivery we have designed a pharmaceutical platform called Bioplex (biological complex). The concept of Bioplex is to link functional entities via hybridising anchors, such as Peptide Nucleic Acids (PNA), directly to naked DNA. In order to promote delivery functional entities consisting of biologically active peptides or carbohydrates, are linked to the PNA anchor. The PNA acts as genetic glue and hybridises with DNA in a sequence specific manner. By using functional entities, which elicit receptor-mediated endocytosis, improved endosomal escape and enhance nuclear entry we wish to improve the transfer of genetic material into the cell. An important aspect is that the functional entities should also have tissue-targeting properties in vivo. Examples of functional entities investigated to date are the Simian virus 40 nuclear localisation signal to improve nuclear uptake and different carbohydrate ligands in order to achieve receptor specific uptake. The delivery system is also endowed with regulatory capability, since the release of functional entities can be controlled. The aim is to create a safe, pharmaceutically defined and stable delivery system for nucleic acids with enhanced transfection properties that can be used in the clinic.

  14. Analysis of edible oil processing options for the BIO-Plex advanced life support system.

    PubMed

    Greenwalt, C J; Hunter, J

    2000-01-01

    Edible oil is a critical component of the proposed plant-based Advanced Life Support (ALS) diet. Soybean, peanut, and single-cell oil are the oil source options to date. In terrestrial manufacture, oil is ordinarily extracted with hexane, an organic solvent. However, exposed solvents are not permitted in the spacecraft environment or in enclosed human tests by National Aeronautics and Space Administration due to their potential danger and handling difficulty. As a result, alternative oil-processing methods will need to be utilized. Preparation and recovery options include traditional dehulling, crushing, conditioning, and flaking, extrusion, pressing, water extraction, and supercritical extraction. These processing options were evaluated on criteria appropriate to the Advanced Life Support System and BIO-Plex application including: product quality, product stability, waste production, risk, energy needs, labor requirements, utilization of nonrenewable resources, usefulness of by-products, and versatility and mass of equipment to determine the most appropriate ALS edible oil-processing operation.

  15. Analysis of edible oil processing options for the BIO-Plex advanced life support system.

    PubMed

    Greenwalt, C J; Hunter, J

    2000-01-01

    Edible oil is a critical component of the proposed plant-based Advanced Life Support (ALS) diet. Soybean, peanut, and single-cell oil are the oil source options to date. In terrestrial manufacture, oil is ordinarily extracted with hexane, an organic solvent. However, exposed solvents are not permitted in the spacecraft environment or in enclosed human tests by National Aeronautics and Space Administration due to their potential danger and handling difficulty. As a result, alternative oil-processing methods will need to be utilized. Preparation and recovery options include traditional dehulling, crushing, conditioning, and flaking, extrusion, pressing, water extraction, and supercritical extraction. These processing options were evaluated on criteria appropriate to the Advanced Life Support System and BIO-Plex application including: product quality, product stability, waste production, risk, energy needs, labor requirements, utilization of nonrenewable resources, usefulness of by-products, and versatility and mass of equipment to determine the most appropriate ALS edible oil-processing operation. PMID:11676438

  16. Analysis of edible oil processing options for the BIO-Plex advanced life support system

    NASA Technical Reports Server (NTRS)

    Greenwalt, C. J.; Hunter, J.

    2000-01-01

    Edible oil is a critical component of the proposed plant-based Advanced Life Support (ALS) diet. Soybean, peanut, and single-cell oil are the oil source options to date. In terrestrial manufacture, oil is ordinarily extracted with hexane, an organic solvent. However, exposed solvents are not permitted in the spacecraft environment or in enclosed human tests by National Aeronautics and Space Administration due to their potential danger and handling difficulty. As a result, alternative oil-processing methods will need to be utilized. Preparation and recovery options include traditional dehulling, crushing, conditioning, and flaking, extrusion, pressing, water extraction, and supercritical extraction. These processing options were evaluated on criteria appropriate to the Advanced Life Support System and BIO-Plex application including: product quality, product stability, waste production, risk, energy needs, labor requirements, utilization of nonrenewable resources, usefulness of by-products, and versatility and mass of equipment to determine the most appropriate ALS edible oil-processing operation.

  17. Kinesin-microtubule interactions during gliding assays under magnetic force

    NASA Astrophysics Data System (ADS)

    Fallesen, Todd L.

    Conventional kinesin is a motor protein capable of converting the chemical energy of ATP into mechanical work. In the cell, this is used to actively transport vesicles through the intracellular matrix. The relationship between the velocity of a single kinesin, as it works against an increasing opposing load, has been well studied. The relationship between the velocity of a cargo being moved by multiple kinesin motors against an opposing load has not been established. A major difficulty in determining the force-velocity relationship for multiple motors is determining the number of motors that are moving a cargo against an opposing load. Here I report on a novel method for detaching microtubules bound to a superparamagnetic bead from kinesin anchor points in an upside down gliding assay using a uniform magnetic field perpendicular to the direction of microtubule travel. The anchor points are presumably kinesin motors bound to the surface which microtubules are gliding over. Determining the distance between anchor points, d, allows the calculation of the average number of kinesins, n, that are moving a microtubule. It is possible to calculate the fraction of motors able to move microtubules as well, which is determined to be ˜ 5%. Using a uniform magnetic field parallel to the direction of microtubule travel, it is possible to impart a uniform magnetic field on a microtubule bound to a superparamagnetic bead. We are able to decrease the average velocity of microtubules driven by multiple kinesin motors moving against an opposing force. Using the average number of kinesins on a microtubule, we estimate that there are an average 2-7 kinesins acting against the opposing force. By fitting Gaussians to the smoothed distributions of microtubule velocities acting against an opposing force, multiple velocities are seen, presumably for n, n-1, n-2, etc motors acting together. When these velocities are scaled for the average number of motors on a microtubule, the force

  18. Bioregenerative Life Support Systems Test Complex (Bio-Plex) Food Processing System: A Dual System

    NASA Technical Reports Server (NTRS)

    Perchonok, Michele; Vittadini, Elena; Peterson, Laurie J.; Swango, Beverly E.; Toerne, Mary E.; Russo, Dane M. (Technical Monitor)

    2001-01-01

    A Bioregenerative Life Support Test Complex, BIO-Plex, is currently being constructed at the Johnson Space Center (JSC) in Houston, TX. This facility will attempt to answer the questions involved in developing a lunar or planetary base. The Food Processing System (FPS) of the BIO-Plex is responsible for supplying food to the crew in coordination with the chosen mission scenario. Long duration space missions require development of both a Transit Food System and of a Lunar or Planetary Food System. These two systems are intrinsically different since the first one will be utilized in the transit vehicle in microgravity conditions with mostly resupplied foods, while the second will be used in conditions of partial gravity (hypogravity) to process foods from crops grown in the facility. The Transit Food System will consist of prepackaged food of extended shelf life. It will be supplemented with salad crops that will be consumed fresh. Microgravity imposes significant limitation on the ability to handle food and allows only for minimal processing. The challenge is to develop food systems similar to the International Space Station or Shuttle Food Systems but with a shelf life of 3 - 5 years. The Lunar or Planetary Food System will allow for food processing of crops due to the presence of some gravitational force (1/6 to 1/3 that of Earth). Crops such as wheat, soybean, rice, potato, peanut, and salad crops, will be processed to final products to provide a nutritious and acceptable diet for the crew. Not only are constraints imposed on the FPS from the crops (e.g., crop variation, availability, storage and shelf-life) but also significant requirements are present for the crew meals (e.g., RDA, high quality, safety, variety). The FPS becomes a fulcrum creating the right connection from crops to crew meals while dealing with issues of integration within a closed self-regenerative system (e.g., safe processing, waste production, volumes, air contaminations, water usage, etc

  19. Biosensors and Bio-Bar Code Assays Based on Biofunctionalized Magnetic Microbeads

    PubMed Central

    Jaffrezic-Renault, Nicole; Martelet, Claude; Chevolot, Yann; Cloarec, Jean-Pierre

    2007-01-01

    This review paper reports the applications of magnetic microbeads in biosensors and bio-bar code assays. Affinity biosensors are presented through different types of transducing systems: electrochemical, piezo electric or magnetic ones, applied to immunodetection and genodetection. Enzymatic biosensors are based on biofunctionalization through magnetic microbeads of a transducer, more often amperometric, potentiometric or conductimetric. The bio-bar code assays relie on a sandwich structure based on specific biological interaction of a magnetic microbead and a nanoparticle with a defined biological molecule. The magnetic particle allows the separation of the reacted target molecules from unreacted ones. The nanoparticles aim at the amplification and the detection of the target molecule. The bio-bar code assays allow the detection at very low concentration of biological molecules, similar to PCR sensitivity.

  20. Lens-free imaging of magnetic particles in DNA assays.

    PubMed

    Colle, Frederik; Vercruysse, Dries; Peeters, Sara; Liu, Chengxun; Stakenborg, Tim; Lagae, Liesbet; Del-Favero, Jurgen

    2013-11-01

    We present a novel opto-magnetic system for the fast and sensitive detection of nucleic acids. The system is based on a lens-free imaging approach resulting in a compact and cheap optical readout of surface hybridized DNA fragments. In our system magnetic particles are attracted towards the detection surface thereby completing the labeling step in less than 1 min. An optimized surface functionalization combined with magnetic manipulation was used to remove all nonspecifically bound magnetic particles from the detection surface. A lens-free image of the specifically bound magnetic particles on the detection surface was recorded by a CMOS imager. This recorded interference pattern was reconstructed in software, to represent the particle image at the focal distance, using little computational power. As a result we were able to detect DNA concentrations down to 10 pM with single particle sensitivity. The possibility of integrated sample preparation by manipulation of magnetic particles, combined with the cheap and highly compact lens-free detection makes our system an ideal candidate for point-of-care diagnostic applications.

  1. Dose-response curve of a microfluidic magnetic bead-based surface coverage sandwich assay.

    PubMed

    Cornaglia, Matteo; Trouillon, Raphaël; Tekin, H Cumhur; Lehnert, Thomas; Gijs, Martin A M

    2015-09-25

    Magnetic micro- and nanoparticles ('magnetic beads') have been used to advantage in many microfluidic devices for sensitive antigen (Ag) detection. Today, assays that use as read-out of the signal the number count of immobilized beads on a surface for quantification of a sample's analyte concentration have been among the most sensitive and have allowed protein detection lower than the fgmL(-1) concentration range. Recently, we have proposed in this category a magnetic bead surface coverage assay (Tekin et al., 2013 [1]), in which 'large' (2.8μm) antibody (Ab)-functionalized magnetic beads captured their Ag from a serum and these Ag-carrying beads were subsequently exposed to a surface pattern of fixed 'small' (1.0μm) Ab-coated magnetic beads. When the system was exposed to a magnetic induction field, the magnet dipole attractive interactions between the two bead types were used as a handle to approach both bead surfaces and assist with Ag-Ab immunocomplex formation, while unspecific binding (in absence of an Ag) of a large bead was reduced by exploiting viscous drag flow. The dose-response curve of this type of assay had two remarkable features: (i) its ability to detect an output signal (i.e. bead number count) for very low Ag concentrations, and (ii) an output signal of the assay that was non-linear with respect to Ag concentration. We explain here the observed dose-response curves and show that the type of interactions and the concept of our assay are in favour of detecting the lowest analyte concentrations (where typically either zero or one Ag is carried per large bead), while higher concentrations are less efficiently detected. We propose a random walk process for the Ag-carrying bead over the magnetic landscape of small beads and this model description explains the enhanced overall capture probability of this assay and its particular non-linear dose response curves.

  2. An Immuno-Magnetic Nanobead Probe Competitive Assay for Rapid Detection of Salmonella choleraesuis.

    PubMed

    Liu, Daofeng; Yu, Zhibiao; Huang, Yanmei; Wang, Shuying; Wang, Jingyun; Guo, Qi; Xu, Chaolian; Xia, Shiqi; Lai, Weihua

    2016-03-01

    A competitive lateral flow assay for the rapid detection of Salmonella choleraesuis was developed. Immuno-magnetic nanobeads were produced by covalently coupling anti-Salmonella choleraesuis antibody to magnetic nanobeads. These immuno-magnetic nanobeads were used as visually detected probes in the subsequent assay. Compared with the traditional sandwich assay, which is used for detecting macro-molecules, this new method was developed based on the competitive relationship between S. choleraesuis in the inspected sample and the outer membrane protein immobilized on the T line. Thus, only one antibody was necessary in the new assay, whereas a pair of rigorously selected antibodies were required in the sandwich assay. The sensitivity of the competitive assay for S. choleraesuis was 1.2 x 10(7) cfu/mL. In addition, no cross reactions were found in the 17 common non-Salmonella bacteria strains and in the 4 Salmonella strains of other serotypes. Thus, with satisfactory sensitivity and specificity, the assay can be applied for the rapid detection of pre-enriched culture that may contain S. choleraesuis. PMID:27455631

  3. An Immuno-Magnetic Nanobead Probe Competitive Assay for Rapid Detection of Salmonella choleraesuis.

    PubMed

    Liu, Daofeng; Yu, Zhibiao; Huang, Yanmei; Wang, Shuying; Wang, Jingyun; Guo, Qi; Xu, Chaolian; Xia, Shiqi; Lai, Weihua

    2016-03-01

    A competitive lateral flow assay for the rapid detection of Salmonella choleraesuis was developed. Immuno-magnetic nanobeads were produced by covalently coupling anti-Salmonella choleraesuis antibody to magnetic nanobeads. These immuno-magnetic nanobeads were used as visually detected probes in the subsequent assay. Compared with the traditional sandwich assay, which is used for detecting macro-molecules, this new method was developed based on the competitive relationship between S. choleraesuis in the inspected sample and the outer membrane protein immobilized on the T line. Thus, only one antibody was necessary in the new assay, whereas a pair of rigorously selected antibodies were required in the sandwich assay. The sensitivity of the competitive assay for S. choleraesuis was 1.2 x 10(7) cfu/mL. In addition, no cross reactions were found in the 17 common non-Salmonella bacteria strains and in the 4 Salmonella strains of other serotypes. Thus, with satisfactory sensitivity and specificity, the assay can be applied for the rapid detection of pre-enriched culture that may contain S. choleraesuis.

  4. Bioregenerative Planetary Life Support Systems Test Complex (BIO-Plex): NASA's Next Human-Rated Testing Facility

    NASA Technical Reports Server (NTRS)

    Tri, Terry O.

    1999-01-01

    As a key component in its ground test bed capability, NASA's Advanced Life Support Program has been developing a large-scale advanced life support test facility capable of supporting long-duration evaluations of integrated bioregenerative life support systems with human test crews. This facility-targeted for evaluation of hypogravity compatible life support systems to be developed for use on planetary surfaces such as Mars or the Moon-is called the Bioregenerative Planetary Life Support Systems Test Complex (BIO-Plex) and is currently under development at the Johnson Space Center. This test bed is comprised of a set of interconnected chambers with a sealed internal environment which are outfitted with systems capable of supporting test crews of four individuals for periods exceeding one year. The advanced technology systems to be tested will consist of both biological and physicochemical components and will perform all required crew life support functions. This presentation provides a description of the proposed test "missions" to be supported by the BIO-Plex and the planned development strategy for the facility.

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

    PubMed

    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

  6. 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

  7. Building bio-assays with magnetic particles on a digital microfluidic platform.

    PubMed

    Kokalj, Tadej; Pérez-Ruiz, Elena; Lammertyn, Jeroen

    2015-09-25

    Digital microfluidics (DMF) has emerged as a promising liquid handling technology for a variety of applications, demonstrating great potential both in terms of miniaturization and automation. DMF is based on the manipulation of discrete, independently controllable liquid droplets, which makes it highly reconfigurable and reprogrammable. One of its most exclusive advantages, compared to microchannel-based microfluidics, is its ability to precisely handle solid nano- and microsized objects, such as magnetic particles. Magnetic particles have become very popular in the last decade, since their high surface-to-volume ratio and the possibility to magnetically separate them from the matrix make them perfect suitable as a solid support for bio-assay development. The potential of magnetic particles in DMF-based bio-assays has been demonstrated for various applications. In this review we discuss the latest developments of magnetic particle-based DMF bio-assays with the aim to present, identify and analyze the trends in the field. We also discuss the state-of-the art of device integration, current status of commercialization and issues that still need to be addressed. With this paper we intend to stimulate researchers to exploit and unveil the potential of these exciting tools, which will shape the future of modern biochemistry, microbiology and biomedical diagnostics. PMID:25813426

  8. Building bio-assays with magnetic particles on a digital microfluidic platform.

    PubMed

    Kokalj, Tadej; Pérez-Ruiz, Elena; Lammertyn, Jeroen

    2015-09-25

    Digital microfluidics (DMF) has emerged as a promising liquid handling technology for a variety of applications, demonstrating great potential both in terms of miniaturization and automation. DMF is based on the manipulation of discrete, independently controllable liquid droplets, which makes it highly reconfigurable and reprogrammable. One of its most exclusive advantages, compared to microchannel-based microfluidics, is its ability to precisely handle solid nano- and microsized objects, such as magnetic particles. Magnetic particles have become very popular in the last decade, since their high surface-to-volume ratio and the possibility to magnetically separate them from the matrix make them perfect suitable as a solid support for bio-assay development. The potential of magnetic particles in DMF-based bio-assays has been demonstrated for various applications. In this review we discuss the latest developments of magnetic particle-based DMF bio-assays with the aim to present, identify and analyze the trends in the field. We also discuss the state-of-the art of device integration, current status of commercialization and issues that still need to be addressed. With this paper we intend to stimulate researchers to exploit and unveil the potential of these exciting tools, which will shape the future of modern biochemistry, microbiology and biomedical diagnostics.

  9. Droplet-based microfluidic washing module for magnetic particle-based assays

    PubMed Central

    Lee, Hun; Xu, Linfeng; Oh, Kwang W.

    2014-01-01

    In this paper, we propose a continuous flow droplet-based microfluidic platform for magnetic particle-based assays by employing in-droplet washing. The droplet-based washing was implemented by traversing functionalized magnetic particles across a laterally merged droplet from one side (containing sample and reagent) to the other (containing buffer) by an external magnetic field. Consequently, the magnetic particles were extracted to a parallel-synchronized train of washing buffer droplets, and unbound reagents were left in an original train of sample droplets. To realize the droplet-based washing function, the following four procedures were sequentially carried in a droplet-based microfluidic device: parallel synchronization of two trains of droplets by using a ladder-like channel network; lateral electrocoalescence by an electric field; magnetic particle manipulation by a magnetic field; and asymmetrical splitting of merged droplets. For the stable droplet synchronization and electrocoalescence, we optimized droplet generation conditions by varying the flow rate ratio (or droplet size). Image analysis was carried out to determine the fluorescent intensity of reagents before and after the washing step. As a result, the unbound reagents in sample droplets were significantly removed by more than a factor of 25 in the single washing step, while the magnetic particles were successfully extracted into washing buffer droplets. As a proof-of-principle, we demonstrate a magnetic particle-based immunoassay with streptavidin-coated magnetic particles and fluorescently labelled biotin in the proposed continuous flow droplet-based microfluidic platform. PMID:25379098

  10. Aptamer-based detection of plasma proteins by an electrochemical assay coupled to magnetic beads.

    PubMed

    Centi, Sonia; Tombelli, Sara; Minunni, Maria; Mascini, Marco

    2007-02-15

    The DNA thrombin aptamer has been extensively investigated, and the coupling of this aptamer to different transduction principles has demonstrated the wide applicability of aptamers as bioreceptors in bioanalytical assays. The goal of this work was to design an aptamer-based sandwich assay with electrochemical detection for thrombin analysis in complex matrixes, using a simple target capturing step by aptamer-functionalized magnetic beads. The conditions for the aptamer immobilization and for the protein binding have been first optimized by surface plasmon resonance, and then transferred to the electrochemical-based assay performed onto screen-printed electrodes. The assay was then applied to the analysis of thrombin in buffer, spiked serum, and plasma and high sensitivity and specificity were found. Moreover, thrombin was generated in situ in plasma by the conversion of its precursor prothrombin, and the formation of thrombin was followed at different times. The concentrations detected by the electrochemical assay were in agreement with a simulation software that mimics the formation of thrombin over time (thrombogram). The proposed work demonstrates that the high specificity of aptamers together with the use of magnetic beads are the key features for aptamer-based analysis in complex matrixes, opening the possibility of a real application to diagnostics or medical investigation.

  11. Switching assay as a novel approach for specific antigen- antibody interaction analysis using magnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Parr, M.; Illarionov, R.; Marchenko, Y.; Yakovleva, L.; Nikolaev, B.; Ischenko, A.; Shevtsov, M.

    2016-08-01

    Switching assay was applied for the detection of antigen-antibody interaction between 70-kDa heat shock protein (Hsp70) and anti-Hsp70 monoclonal antibodies in water solutions using conjugates with magnetic iron oxide nanoparticles (MNPs). Hsp70 is a ubiquitous intracellular protein that plays a crucial role in cancerogenesis and many other pathologies. Detection of the Hsp70 level in the biological fluids might have a prognostic and diagnostic value in clinic. The developed switch assay for the detection of Hsp70 demonstrated high sensitivity for antigen-antibody interaction analysis thus proving its potential for further preclinical and clinical studies.

  12. Magnetic enzyme-linked immunosorbent assay (MELISA) for determination of specific IgG in paracoccidioidomycosis.

    PubMed

    de Camargo, Z P; Guesdon, J L; Drouhet, E; Improvisi, L

    1984-01-01

    A magnetic solid phase enzyme-linked immunosorbent assay (MELISA) for quantification of IgG antibodies to somatic and metabolic antigens of Paracoccidioides brasiliensis was developed. Activation of magnetic polyacrylamide agarose beads with concanavalin A was superior to glutaraldehyde activation, and test sensitivity was higher for somatic than for metabolic antigens. Comparative MELISA, counterimmunoelectrophoresis and erythroimmunoassay tests with sera from 33 proven cases of paracoccidioidomycosis, 14 cases of histoplasmosis and 20 normal human sera showed the MELISA could distinguish antibody levels in paracoccidioidomycosis from those in normal sera; however two sera from histoplasmosis cases cross-reacted in the MELISA. MELISA is a rapid test (5-6 h) and the results suggest it has considerable potential value for assay of anti-P. brasiliensis antibodies. PMID:6438813

  13. Stability study for magnetic reagent assaying Hb and HbA1c

    NASA Astrophysics Data System (ADS)

    Hsieh, Wen-Pin; Chieh, J. J.; Yang, C. C.; Yang, S. Y.; Chen, Po-Yu; Huang, Yu-Hao; Hong, Y. W.; Horng, H. E.

    2013-01-01

    Reagents for magnetically labeled immunoassay on human Hb and human HbA1c have been synthesized. The reagents consist of Fe3O4 magnetic particles biofunctionalized with antibodies against Hb and HbA1c. It has been demonstrated that the reagents can be applied to quantitatively detect Hb and HbA1c by using immunomagnetic reduction assay. In addition to characterizing the assay properties, such as the standard curve and the low-detection limit, the stability of reagents is investigated. To do this, the temporal dependence of particle sizes and the bio-activity of reagents are monitored. The results show that the reagents are highly stable when stored at 2-8 °C. This means that the reagents synthesized in this work are promising for practical applications.

  14. A Magnetic Bead-Based Protein Kinase Assay with Dual Detection Techniques

    PubMed Central

    Zhou, Guangchang; Sylvester, Juliesta E.; Wu, Ding; Veach, Darren R.; Kron, Stephen J.

    2010-01-01

    A novel magnetic bead-based protein kinase assay was developed using MALDI-TOF mass spectrometry (MALDI-TOF MS) and immuno-chemifluorescence as two independent detection techniques. Abltide substrate was immobilized onto magnetic beads via non-covalent biotin-streptavidin interactions. This non-covalent immobilization strategy facilitated peptide release and allowed MALDI-TOF MS analysis of substrate phosphorylation. The use of magnetic beads provided rapid sample handling and allowed secondary analysis by immuno-chemifluorescence to determine the degree of substrate phosphorylation. This dual detection technique was used to evaluate the inhibition of c-Abl kinase by imatinib and dasatinib. For each inhibitor, IC50 (half-maximal inhibitory concentration) values determined by these two different detection methods were consistent and close to values reported in the literature. The high-throughput potential of this new approach to kinase assays was preliminarily demonstrated by screening a chemical library consisting of 31 compounds against c-Abl kinase using a 96-well plate. In this proof-of-principle experiment, both MALDI-TOF MS and immuno-chemifluorescence were able to compare inhibitor potencies with consistent values. Dual detection may significantly enhance the reliability of chemical library screening and identify false positives and negatives. Formatted for 96-well plates and with high-throughput potential, this dual detection kinase assay may provide a rapid, reliable and inexpensive route to the discovery of small molecule drug leads. PMID:20807497

  15. The clinical utility of anti-chromatin antibodies as measured by BioPlex 2200 in the diagnosis of systemic lupus erythematosus versus other rheumatic diseases

    PubMed Central

    Bose, Nilanjana; Wang, Xiaofeng; Gupta, Majula; Yao, Qingping

    2012-01-01

    Background The detection of autoantibodies is indispensable to systemic lupus erythematosus (SLE). Bioplex 2200 ANA screen is a multiplex immunoassay system that allows simultaneous determination of autoantibodies to extractable nuclear antigens (ENA) including anti-chromatin antibodies (ACAs). However, the clinical significance of the ACAs by this new method in SLE patients has not been studied in comparison with other rheumatic disorders. We performed a retrospective study of patients with rheumatic diseases to assess the diagnostic value of the ACAs by Bioplex 2200 method in SLE. Methods Adult patients with rheumatic complaints seen by rheumatologists at the Cleveland Clinic between January 2008 and February 2010 were screened for positive anti-ENA antibodies by the Bioplex 2200. Patients with positive anti-ENA antibodies were classified into two populations based upon the presence and absence of the ACAs. We retrospectively studied the clinical and laboratory data of these patients. Results A total of 764 subjects with positive anti-ENA antibodies were screened, including 115 with positive ACAs. There were 93 SLE patients consisting of 58 with positive ACAs and 35 with negative ACAs. The sensitivity, specificity, positive predictive value and negative predictive value of the ACAs in SLE were 62.4%, 91.5%, 50.4% and 94.6% respectively. Apart from SLE, positive ACAs were associated with mixed connective tissue disease (MCTD)/undifferentiated CTD (UCTD) and other autoimmune diseases. No correlation was found between the ACAs and lupus glomerulonephritis or anti-dsDNA antibodies. Conclusions Measurement of the ACAs by the Bioplex 2200 is specific for diagnosing SLE but not useful for differentiating between SLE and MCTD/UCTD. PMID:22993651

  16. Magnetic bead-quantum dot assay for detection of a biomarker for traumatic brain injury

    NASA Astrophysics Data System (ADS)

    Kim, Chloe; Searson, Peter C.

    2015-10-01

    Current diagnostic methods for traumatic brain injury (TBI), which accounts for 15% of all emergency room visits, are limited to neuroimaging modalities. The challenges of accurate diagnosis and monitoring of TBI have created the need for a simple and sensitive blood test to detect brain-specific biomarkers. Here we report on an assay for detection of S100B, a putative biomarker for TBI, using antibody-conjugated magnetic beads for capture of the protein, and antibody-conjugated quantum dots for optical detection. From Western Blot, we show efficient antigen capture and concentration by the magnetic beads. Using magnetic bead capture and quantum dot detection in serum samples, we show a wide detection range and detection limit below the clinical cut-off level.Current diagnostic methods for traumatic brain injury (TBI), which accounts for 15% of all emergency room visits, are limited to neuroimaging modalities. The challenges of accurate diagnosis and monitoring of TBI have created the need for a simple and sensitive blood test to detect brain-specific biomarkers. Here we report on an assay for detection of S100B, a putative biomarker for TBI, using antibody-conjugated magnetic beads for capture of the protein, and antibody-conjugated quantum dots for optical detection. From Western Blot, we show efficient antigen capture and concentration by the magnetic beads. Using magnetic bead capture and quantum dot detection in serum samples, we show a wide detection range and detection limit below the clinical cut-off level. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr05608j

  17. Design criteria for developing low-resource magnetic bead assays using surface tension valves.

    PubMed

    Adams, Nicholas M; Creecy, Amy E; Majors, Catherine E; Wariso, Bathsheba A; Short, Philip A; Wright, David W; Haselton, Frederick R

    2013-01-01

    Many assays for biological sample processing and diagnostics are not suitable for use in settings that lack laboratory resources. We have recently described a simple, self-contained format based on magnetic beads for extracting infectious disease biomarkers from complex biological samples, which significantly reduces the time, expertise, and infrastructure required. This self-contained format has the potential to facilitate the application of other laboratory-based sample processing assays in low-resource settings. The technology is enabled by immiscible fluid barriers, or surface tension valves, which stably separate adjacent processing solutions within millimeter-diameter tubing and simultaneously permit the transit of magnetic beads across the interfaces. In this report, we identify the physical parameters of the materials that maximize fluid stability and bead transport and minimize solution carryover. We found that fluid stability is maximized with ≤0.8 mm i.d. tubing, valve fluids of similar density to the adjacent solutions, and tubing with ≤20 dyn/cm surface energy. Maximizing bead transport was achieved using ≥2.4 mm i.d. tubing, mineral oil valve fluid, and a mass of 1-3 mg beads. The amount of solution carryover across a surface tension valve was minimized using ≤0.2 mg of beads, tubing with ≤20 dyn/cm surface energy, and air separators. The most favorable parameter space for valve stability and bead transport was identified by combining our experimental results into a single plot using two dimensionless numbers. A strategy is presented for developing additional self-contained assays based on magnetic beads and surface tension valves for low-resource diagnostic applications. PMID:24403996

  18. Design criteria for developing low-resource magnetic bead assays using surface tension valves

    PubMed Central

    Adams, Nicholas M.; Creecy, Amy E.; Majors, Catherine E.; Wariso, Bathsheba A.; Short, Philip A.; Wright, David W.; Haselton, Frederick R.

    2013-01-01

    Many assays for biological sample processing and diagnostics are not suitable for use in settings that lack laboratory resources. We have recently described a simple, self-contained format based on magnetic beads for extracting infectious disease biomarkers from complex biological samples, which significantly reduces the time, expertise, and infrastructure required. This self-contained format has the potential to facilitate the application of other laboratory-based sample processing assays in low-resource settings. The technology is enabled by immiscible fluid barriers, or surface tension valves, which stably separate adjacent processing solutions within millimeter-diameter tubing and simultaneously permit the transit of magnetic beads across the interfaces. In this report, we identify the physical parameters of the materials that maximize fluid stability and bead transport and minimize solution carryover. We found that fluid stability is maximized with ≤0.8 mm i.d. tubing, valve fluids of similar density to the adjacent solutions, and tubing with ≤20 dyn/cm surface energy. Maximizing bead transport was achieved using ≥2.4 mm i.d. tubing, mineral oil valve fluid, and a mass of 1-3 mg beads. The amount of solution carryover across a surface tension valve was minimized using ≤0.2 mg of beads, tubing with ≤20 dyn/cm surface energy, and air separators. The most favorable parameter space for valve stability and bead transport was identified by combining our experimental results into a single plot using two dimensionless numbers. A strategy is presented for developing additional self-contained assays based on magnetic beads and surface tension valves for low-resource diagnostic applications. PMID:24403996

  19. Different approaches for the detection of thrombin by an electrochemical aptamer-based assay coupled to magnetic beads.

    PubMed

    Centi, S; Messina, G; Tombelli, S; Palchetti, I; Mascini, M

    2008-06-15

    Different assay formats based on the coupling of magnetic beads with electrochemical transduction were compared here for the detection of thrombin by using a thrombin specific aptamer. By using the thrombin-binding aptamer, a direct and an indirect competitive assay for thrombin have been developed by immobilising the aptamer or the protein, respectively. Moreover, another strategy was based on the direct measurement of the enzymatic product of thrombin captured by the immobilised aptamer. All the assays were developed by coupling the electrochemical transduction with the innovative and advantageous use of magnetic beads. The assays based on the immobilisation of the protein were not successful since no binding was recorded between thrombin and its aptamer. With the direct competitive assay, when the aptamer was immobilised onto the magnetic beads, a detection limit of 430nM for thrombin was achieved. A lower detection limit for the protein (175nM) was instead obtained by detecting the product of the enzymatic reaction catalysed by thrombin. All these assays were finally compared with a sandwich assay which reached a detection limit of 0.45nM of thrombin demonstrating the best analytical performances. With this comparison the importance of a deep study on the different analytical approaches for thrombin detection to reach the performances of the best assay configuration has been demonstrated.

  20. A highly sensitive and flexible magnetic nanoprobe labeled immunochromatographic assay platform for pathogen Vibrio parahaemolyticus.

    PubMed

    Liu, Yingying; Zhang, Zhaohuan; Wang, Yilong; Zhao, Yong; Lu, Ying; Xu, Xiaowei; Yan, Jun; Pan, Yingjie

    2015-10-15

    A magnetic nanoprobe labeled immunochromatographic test strip (MNP/ICTS) was developed to detect food-borne pathogen Vibrio parahaemolyticus. Specific antibody against V. parahaemolyticus was used as test line by coating onto the nitrocellulose membrane. Magnetic nanoprobe was prepared by immobilizing the specific antibody onto the surface of superparamagnetic nanoparticles. Specificity and sensitivity of the MNP/ICTS system were verified by artificially contaminated shrimp homogenate samples. Reliability and application feasibility of the MNP/ICTS system were demonstrated by using seafood samples (n=36). Comparing with polymerase chain reaction (PCR) and traditional culture methods, the MNP/ICTS system is found to be not only a rapid qualitative analysis (~10 min), but also an accurately quantitative detection platform. Through its rapid magnetic separation property, the MNP/ICTS system is capable to flexibly combine with a sample enrichment and pre-incubation process. This combination makes the qualitative sensitivity for the food samples surged more than 100-fold. A naked-eye observation of 1.58×10(2) CFU/g V. parahaemolyticus was realized. This sensitivity could meet the V. parahaemolyticus test threshold value in many countries. Also, the total sample pre-treatment plus MNP/ICTS assay only needs about 4.5h. Namely, we can get test results in a day. Hence, the developed MNP/ICTS assay platform is simple, rapid and highly sensitive. It is a flexible test platform for pathogen detection. The favorable comparison with PCR and culture methods further proves that the developed MNP/ICTS is applicable into food-borne pathogen or other areas where a simple, rapid, sensitive and point-of-care analysis is desirable. PMID:26188497

  1. A highly sensitive and flexible magnetic nanoprobe labeled immunochromatographic assay platform for pathogen Vibrio parahaemolyticus.

    PubMed

    Liu, Yingying; Zhang, Zhaohuan; Wang, Yilong; Zhao, Yong; Lu, Ying; Xu, Xiaowei; Yan, Jun; Pan, Yingjie

    2015-10-15

    A magnetic nanoprobe labeled immunochromatographic test strip (MNP/ICTS) was developed to detect food-borne pathogen Vibrio parahaemolyticus. Specific antibody against V. parahaemolyticus was used as test line by coating onto the nitrocellulose membrane. Magnetic nanoprobe was prepared by immobilizing the specific antibody onto the surface of superparamagnetic nanoparticles. Specificity and sensitivity of the MNP/ICTS system were verified by artificially contaminated shrimp homogenate samples. Reliability and application feasibility of the MNP/ICTS system were demonstrated by using seafood samples (n=36). Comparing with polymerase chain reaction (PCR) and traditional culture methods, the MNP/ICTS system is found to be not only a rapid qualitative analysis (~10 min), but also an accurately quantitative detection platform. Through its rapid magnetic separation property, the MNP/ICTS system is capable to flexibly combine with a sample enrichment and pre-incubation process. This combination makes the qualitative sensitivity for the food samples surged more than 100-fold. A naked-eye observation of 1.58×10(2) CFU/g V. parahaemolyticus was realized. This sensitivity could meet the V. parahaemolyticus test threshold value in many countries. Also, the total sample pre-treatment plus MNP/ICTS assay only needs about 4.5h. Namely, we can get test results in a day. Hence, the developed MNP/ICTS assay platform is simple, rapid and highly sensitive. It is a flexible test platform for pathogen detection. The favorable comparison with PCR and culture methods further proves that the developed MNP/ICTS is applicable into food-borne pathogen or other areas where a simple, rapid, sensitive and point-of-care analysis is desirable.

  2. Magnetic zirconium hexacyanoferrate(II) nanoparticle as tracing tag for electrochemical DNA assay.

    PubMed

    Zhang, Guang-Yao; Deng, Sheng-Yuan; Cai, Wen-Rong; Cosnier, Serge; Zhang, Xue-Ji; Shan, Dan

    2015-09-01

    Novel multifunctional magnetic zirconium hexacyanoferrate nanoparticles (ZrHCF MNPs) were prepared, which consisted of magnetic beads (MBs) inner core and zirconium hexacyanoferrate(II) (ZrHCF) outer shell. As an artificial peroxidase, the ZrHCF MNPs exhibited remarkable electrocatalytic properties in the reduction of H2O2 at 0.2 V vs saturated calomel electrode (SCE). On the basis of the bonding interaction between Zr (IV) of the shell ZrHCF framework and phosphonate groups, the 5'-phosphorylated ssDNA probes with a consecutive stretch of guanines as a spacer could be incorporated in ZrHCF MNPs easily. Thus, DNA-grafted ZrHCF MNPs could be simply obtained by magnetic separation. The prepared nanoelectrocatalyst was further used as signal nanoprobe for the ultrasensitive electrochemical DNA assay. Under optimal conditions, the proposed biosensor presents high sensitivity for detecting target DNA with a linear range from 1.0 fM to 1.0 nM and a low detection limit of 0.43 fM. Moreover, it exhibits good performance with excellent selectivity, high stability, and acceptable fabrication reproducibility.

  3. Indicator-based and indicator-free magnetic assays connected with disposable electrochemical nucleic acid sensor system.

    PubMed

    Karadeniz, Hakan; Erdem, Arzum; Kuralay, Filiz; Jelen, Frantisek

    2009-04-15

    An indicator-based and indicator-free magnetic assays connected with a disposable pencil graphite electrode (PGE) were successfully developed, and also compared for the electrochemical detection of DNA hybridization. The oxidation signals of echinomycin (ECHI) and electroactive DNA bases, guanine and adenine, respectively were monitored in the presence of DNA hybridization by using differential pulse voltammetry (DPV) technique. The biotinylated probe was immobilized onto the magnetic beads (magnetic particles, microspheres) and hybridization with its complementary target at the surface of particles within the medium was exhibited successfully using electrochemical sensor system. For the selectivity studies, the results represent that both indicator-based and indicator-free magnetic assays provide a better discrimination for DNA hybridization compared to duplex with one-base or more mismatches. The detection limits (S/N=3) of the magnetic assays based on indicator or indicator-free were found in nM concentration level of target using disposable sensor technology with good reproducibility. The characterization and advantages of both proposed magnetic assays connected with a disposable electrochemical sensor are also discussed and compared with those methods previously reported in the literature.

  4. Giant Magnetoresistive Sensors and Magnetic Labels for Chip-Scale Detection of Immunosorbent Assays

    SciTech Connect

    Millen, Rachel Lora

    2005-01-01

    The combination of giant magnetoresistive sensors, magnetic labeling strategies, and biomolecule detection is just beginning to be explored. New readout methods and assay formats are necessary for biomolecules detection to flourish. The work presented in this dissertation describes steps toward the creation of a novel detection method for bioassays utilizing giant magnetoresistive sensors as the readout method. The introduction section contains a brief review of some of the current methods of bioassay readout. The theoretical underpinnings of the giant magnetoresistive effect are also discussed. Finally, the more prominent types of giant magnetoresistive sensors are described, as well as their complicated fabrication. Four data chapters follow the introduction; each chapter is presented as a separate manuscript, either already published or soon to be submitted. Chapter 1 presents research efforts toward the production of a bioassay on the surface of a gold-modified GMR sensor. The testing of this methodology involved the capture of goat a-mouse-coated magnetic nanoparticles on the mouse IgG-modified gold surface. The second, third and fourth chapters describe the utilization of a self-referenced sample stick for scanning across the GMR sensor. The sample stick consisted of alternating magnetic reference and bioactive gold addresses. Chapter 2 is concerned with the characterization of both the scanning readout method and the binding and detection of streptavidin-coated magnetic particles to a biotinylated surface. Chapter 3 advances the sample stick readout with the use of the system for detection of a sandwich immunoassay with rabbit IgG proteins. Finally, simultaneous detection of three IgG proteins is demonstrated in Chapter 4. The dissertation is concluded with a brief summary of the research presented and a discussion of the possible future applications and direction of this work.

  5. Fluorine nuclear magnetic resonance-based assay in living mammalian cells.

    PubMed

    Veronesi, Marina; Giacomina, Francesca; Romeo, Elisa; Castellani, Beatrice; Ottonello, Giuliana; Lambruschini, Chiara; Garau, Gianpiero; Scarpelli, Rita; Bandiera, Tiziano; Piomelli, Daniele; Dalvit, Claudio

    2016-02-15

    Nuclear magnetic resonance (NMR)-based screening has been recognized as a powerful approach for the identification and characterization of molecules interacting with pharmaceutical targets. Indeed, several NMR methods have been developed and successfully applied to many drug discovery projects. Whereas most of these approaches have targeted isolated biomolecular receptors, very few cases are reported with the screening performed in intact cells and cell extracts. Here we report the first successful application of the fluorine NMR-based assay n-FABS (n-fluorine atoms for biochemical screening) in living mammalian cells expressing the membrane protein fatty acid amide hydrolase (FAAH). This method allows the identification of both weak and potent inhibitors and the measurement of their potency in a physiological environment.

  6. NMR-DMF: a modular nuclear magnetic resonance-digital microfluidics system for biological assays.

    PubMed

    Lei, Ka-Meng; Mak, Pui-In; Law, Man-Kay; Martins, Rui P

    2014-12-01

    We present a modular nuclear magnetic resonance-digital microfluidics (NMR-DMF) system as a portable diagnostic platform for miniaturized biological assays. With increasing number of combinations between designed probes and a specific target, NMR has become an accurate and rapid assay tool, which is capable of detecting particular kinds of proteins, DNAs, bacteria and cells with a customized probe quantitatively. Traditional sample operation (e.g., manipulation and mixing) relied heavily on human efforts. We herein propose a modular NMR-DMF system to allow the electronic automation of multi-step reaction-screening protocols. A figure-8 shaped coil is proposed to enlarge the usable inner space of a portable magnet by 4.16 times, generating a radio frequency (RF) excitation field in the planar direction. By electronically managing the electro-wetting-on-dielectric (EWOD) effects over an electrode array, preloaded droplets with the inclusion of biological constituents and targets can be programmed to mix and be guided to the detection site (3.5 × 3.5 mm(2)) for high-sensitivity NMR screening (static B field: 0.46 T, RF field: 1.43 mT per ampere), with the result (voltage signal) displayed in real-time. To show the system's utility, automated real-time identification of 100 pM of avidin in a 14 μL droplet was achieved. The system shows promise as a robust and portable diagnostic device for a wide variety of biological analyses and screening applications. PMID:25315808

  7. NMR-DMF: a modular nuclear magnetic resonance-digital microfluidics system for biological assays.

    PubMed

    Lei, Ka-Meng; Mak, Pui-In; Law, Man-Kay; Martins, Rui P

    2014-12-01

    We present a modular nuclear magnetic resonance-digital microfluidics (NMR-DMF) system as a portable diagnostic platform for miniaturized biological assays. With increasing number of combinations between designed probes and a specific target, NMR has become an accurate and rapid assay tool, which is capable of detecting particular kinds of proteins, DNAs, bacteria and cells with a customized probe quantitatively. Traditional sample operation (e.g., manipulation and mixing) relied heavily on human efforts. We herein propose a modular NMR-DMF system to allow the electronic automation of multi-step reaction-screening protocols. A figure-8 shaped coil is proposed to enlarge the usable inner space of a portable magnet by 4.16 times, generating a radio frequency (RF) excitation field in the planar direction. By electronically managing the electro-wetting-on-dielectric (EWOD) effects over an electrode array, preloaded droplets with the inclusion of biological constituents and targets can be programmed to mix and be guided to the detection site (3.5 × 3.5 mm(2)) for high-sensitivity NMR screening (static B field: 0.46 T, RF field: 1.43 mT per ampere), with the result (voltage signal) displayed in real-time. To show the system's utility, automated real-time identification of 100 pM of avidin in a 14 μL droplet was achieved. The system shows promise as a robust and portable diagnostic device for a wide variety of biological analyses and screening applications.

  8. A low cost and high throughput magnetic bead-based immuno-agglutination assay in confined droplets.

    PubMed

    Teste, Bruno; Ali-Cherif, Anaïs; Viovy, Jean Louis; Malaquin, Laurent

    2013-06-21

    Although passive immuno-agglutination assays consist of one step and simple procedures, they are usually not adapted for high throughput analyses and they require expensive and bulky equipment for quantitation steps. Here we demonstrate a low cost, multimodal and high throughput immuno-agglutination assay that relies on a combination of magnetic beads (MBs), droplets microfluidics and magnetic tweezers. Antibody coated MBs were used as a capture support in the homogeneous phase. Following the immune interaction, water in oil droplets containing MBs and analytes were generated and transported in Teflon tubing. When passing in between magnetic tweezers, the MBs contained in the droplets were magnetically confined in order to enhance the agglutination rate and kinetics. When releasing the magnetic field, the internal recirculation flows in the droplet induce shear forces that favor MBs redispersion. In the presence of the analyte, the system preserves specific interactions and MBs stay in the aggregated state while in the case of a non-specific analyte, redispersion of particles occurs. The analyte quantitation procedure relies on the MBs redispersion rate within the droplet. The influence of different parameters such as magnetic field intensity, flow rate and MBs concentration on the agglutination performances have been investigated and optimized. Although the immuno-agglutination assay described in this work may not compete with enzyme linked immunosorbent assay (ELISA) in terms of sensitivity, it offers major advantages regarding the reagents consumption (analysis is performed in sub microliter droplet) and the platform cost that yields to very cheap analyses. Moreover the fully automated analysis procedure provides reproducible analyses with throughput well above those of existing technologies. We demonstrated the detection of biotinylated phosphatase alkaline in 100 nL sample volumes with an analysis rate of 300 assays per hour and a limit of detection of 100 pM.

  9. Ionic strength assay via polyacrylate-ferriferrous oxide magnetic photonic crystals.

    PubMed

    Li, Yan-Ran; Sun, Ye; Wang, He-Fang

    2015-05-21

    Convenient reading out and/or determination of ionic strength (IS) is of great significance for both scientific research and real life applications. We presented here a novel method for the rapid and sensitive IS assay based on the electrolyte-induced sensitive wavelength blueshifts of the reflection spectra of polyacrylate capped Fe3O4 magnetic photonic crystals (PA-Fe3O4-MPCs). For HCl, MgSO4 and the common electrolytes corresponding to the salinity of seawater (including NaCl, KCl, MgCl2, CaCl2, Na2SO4 and their mixtures), the PA-Fe3O4-MPCs displayed wavelength blueshifts identical to the total IS of the aqueous solutions, regardless of the kind of above-mentioned electrolytes in the solutions. Besides, the PA-Fe3O4-MPCs exhibited relatively high sensitivity (an average of 294 nm L mmol(-1) in the range of 0.05-0.30 mmol L(-1), and an even higher value of 386 nm L mmol(-1) at 0.05-0.15 mmol L(-1)) and fast response (within 8 s) to the IS of aqueous solutions. The relative standard deviation (RSD) for IS (NaCl, 0.1 mmol L(-1)) was 4.4% (n = 5). The developed method was applied to determine the salinity of seawater samples, and the determined results were validated by the traditional standard chlorinity titration and electric conductimetry method. The recoveries were in the range of 92-104%. The proposed PA-Fe3O4-MPCs based reflectometry method would have great potential for IS and salinity assays.

  10. A rapid assay for Hendra virus IgG antibody detection and its titre estimation using magnetic nanoparticles and phycoerythrin.

    PubMed

    Gao, Yuan; Pallister, Jackie; Lapierre, Florian; Crameri, Gary; Wang, Lin-Fa; Zhu, Yonggang

    2015-09-15

    Detection of Hendra viral IgG antibody in animal sera is useful for surveillance following a virus outbreak. The commonly used enzyme-linked immunosorbent assay and fluorescence-based Luminex assay typically consist of three steps and take at least several hours to complete. We have simplified the procedure to two steps in an effort to develop a rapid procedure for IgG antibody, but not IgM antibody, detection. This is achieved by conjugating the fluorescence label R-phycoerythrin directly onto the IgG binding protein Protein G. The use of magnetic nanoparticles, due to their large specific surface area, has helped reduce each of the binding steps to 20 min. As a result, the whole assay can be completed in 60 min. We also demonstrate a method to quickly estimate IgG antibody titres by assaying the sera at only two dilutions (i.e. 1:20 and 1:1000) and using the fluorescence ratio at these dilutions as an indicator of antibody titre. The results of this approach correlated well with the well-regarded serum neutralization test in virus antibody assays. This protocol reported here can be adopted in Luminex assays, fluorescence-linked immunosorbent assays and assays on microfluidics platforms for rapid antibody surveillance of Hendra and other viruses.

  11. A rapid assay for Hendra virus IgG antibody detection and its titre estimation using magnetic nanoparticles and phycoerythrin.

    PubMed

    Gao, Yuan; Pallister, Jackie; Lapierre, Florian; Crameri, Gary; Wang, Lin-Fa; Zhu, Yonggang

    2015-09-15

    Detection of Hendra viral IgG antibody in animal sera is useful for surveillance following a virus outbreak. The commonly used enzyme-linked immunosorbent assay and fluorescence-based Luminex assay typically consist of three steps and take at least several hours to complete. We have simplified the procedure to two steps in an effort to develop a rapid procedure for IgG antibody, but not IgM antibody, detection. This is achieved by conjugating the fluorescence label R-phycoerythrin directly onto the IgG binding protein Protein G. The use of magnetic nanoparticles, due to their large specific surface area, has helped reduce each of the binding steps to 20 min. As a result, the whole assay can be completed in 60 min. We also demonstrate a method to quickly estimate IgG antibody titres by assaying the sera at only two dilutions (i.e. 1:20 and 1:1000) and using the fluorescence ratio at these dilutions as an indicator of antibody titre. The results of this approach correlated well with the well-regarded serum neutralization test in virus antibody assays. This protocol reported here can be adopted in Luminex assays, fluorescence-linked immunosorbent assays and assays on microfluidics platforms for rapid antibody surveillance of Hendra and other viruses. PMID:26141730

  12. Preliminary Development of a DNA Aptamer-Magnetic Bead Capture Electrochemiluminescence Sandwich Assay for Brain Natriuretic Peptide

    PubMed Central

    Bruno, John G.; Richarte, Alicia M.; Phillips, Taylor

    2014-01-01

    Fifty-two candidate DNA aptamer sequences were selected for binding to the cardiovascular biomarker B-type or brain natriuretic peptide (BNP). Candidate aptamers were screened to rank their relative affinities against BNP by an aptamer-based ELISA-like aptamer microplate assay (ELASA). The highest affinity aptamers from ELASA screening were also paired in all possible combinations and screened for electrochemiluminescence (ECL) assay potential in capture aptamer-magnetic bead and ruthenium trisbipyridine (Ru(bpy)32+)-reporter aptamer sandwich formats. The top ECL sandwich combinations utilized the same aptamer pair in either capture or reporting roles with nanogram to low picogram per mL levels of detection even in 50% human serum. ECL assay sensitivity and linearity even in 50% human serum suggest that the aptamer-based assay is at least comparable to other reported immunoassays for BNP. PMID:24764602

  13. Magnetic nanoparticle based purification and enzyme-linked immunosorbent assay using monoclonal antibody against enrofloxacin

    PubMed Central

    Kim, Nam-Gun; Kim, Myeong-Ae; Park, Young-Il; Jung, Tae-Sung; Son, Seong-Wan; So, ByungJae

    2015-01-01

    Monoclonal anti-enrofloxacin antibody was prepared for a direct competitive enzyme-linked immunosorbent assay (ELISA) and purification system using monoclonal antibody (mAb) coupled magnetic nanoparticles (MNPs). The IC50 values of the developed mAb for enrofloxacin (ENR), ciprofloxacin, difloxacin, sarafloxacin, pefloxacin, and norfloxacin were 5.0, 8.3, 9.7, 21.7, 36.0, and 63.7 ng/mL, respectively. The lowest detectable level of ENR was 0.7 ng/mL in the prepared ELISA system. To validate the developed ELISA in the food matrix, known amounts of ENR were spiked in meat and egg samples at 10, 20 and 30 ng/mL. Recoveries for ENR ranged from 72.9 to 113.16% with a coefficient of variation (CV) of 2.42 to 10.11%. The applicability of the mAb-MNP system was verified by testing the recoveries for ENR residue in three different matrices. Recoveries for ENR ranged from 75.16 to 86.36%, while the CV ranged from 5.08 to 11.53%. Overall, ENR-specific monoclonal antibody was prepared and developed for use in competitive to ELISAs for the detection of ENR in animal meat samples. Furthermore, we suggest that a purification system for ENR using mAb-coupled MNPs could be useful for determination of ENR residue in food. PMID:26040610

  14. Colorimetric Glucose Assay Based on Magnetic Particles Having Pseudo-peroxidase Activity and Immobilized Glucose Oxidase.

    PubMed

    Martinkova, Pavla; Opatrilova, Radka; Kruzliak, Peter; Styriak, Igor; Pohanka, Miroslav

    2016-05-01

    Magnetic particles (MPs) are currently used as a suitable alternative for peroxidase in the construction of novel biosensors, analytic and diagnostic methods. Their better chemical and thermal stabilities predestine them as appropriate pseudo-enzymatic catalysts. In this point of view, our research was focused on preparation of simply and fast method for immobilization of glucose oxidase onto surface of MPs with peroxidase-like activity. Spectrophotometric method (wavelength 450 nm) optimized for glucose determination using modified MPs has been successfully developed. Concentration curve for optimization of method was assayed, and Michaelis-Menten constant (K m) calculated, maximum reaction rate (V max), limit of detection, and correlation coefficient were determined to be 0.13 mmol/l (2.34 mg/dl), 1.79 pkat, 3.74 µmol/l (0.067 mg/dl), and 0.996, respectively. Interferences of other sugars such as sucrose, sorbitol, deoxyribose, maltose, and fructose were determined as well as effect of substances presenting in plasma (ascorbic acid, reduced glutathione, trolox, and urea). Results in comparison with positive and negative controls showed no interferences of the other sugars and no influence of plasma substances to measuring of glucose. The constructed method showed corresponding results with linear dependence and a correlation coefficient of 0.997. Possibility of repeated use of modified MPs was successfully proved. PMID:27041274

  15. Tandem assays of protein and glucose with functionalized core/shell particles based on magnetic separation and surface-enhanced Raman scattering.

    PubMed

    Kong, Xianming; Yu, Qian; Lv, Zhongpeng; Du, Xuezhong

    2013-10-11

    Tandem assays of protein and glucose in combination with mannose-functionalized Fe3 O4 @SiO2 and Ag@SiO2 tag particles have promising potential in effective magnetic separation and highly sensitive and selective SERS assays of biomaterials. It is for the first time that tandem assay of glucose is developed using SERS based on the Con A-sandwiched microstructures between the functionalized magnetic and tag particles.

  16. A liquid phase affinity capture assay using magnetic beads to study protein-protein interaction: the poliovirus-nanobody example.

    PubMed

    Schotte, Lise; Rombaut, Bart; Thys, Bert

    2012-05-29

    In this article, a simple, quantitative, liquid phase affinity capture assay is presented. Provided that one protein can be tagged and another protein labeled, this method can be implemented for the investigation of protein-protein interactions. It is based on one hand on the recognition of the tagged protein by cobalt coated magnetic beads and on the other hand on the interaction between the tagged protein and a second specific protein that is labeled. First, the labeled and tagged proteins are mixed and incubated at room temperature. The magnetic beads, that recognize the tag, are added and the bound fraction of labeled protein is separated from the unbound fraction using magnets. The amount of labeled protein that is captured can be determined in an indirect way by measuring the signal of the labeled protein remained in the unbound fraction. The described liquid phase affinity assay is extremely useful when conformational conversion sensitive proteins are assayed. The development and application of the assay is demonstrated for the interaction between poliovirus and poliovirus recognizing nanobodies(1). Since poliovirus is sensitive to conformational conversion(2) when attached to a solid surface (unpublished results), the use of ELISA is limited and a liquid phase based system should therefore be preferred. An example of a liquid phase based system often used in polioresearch(3,4) is the micro protein A-immunoprecipitation test(5). Even though this test has proven its applicability, it requires an Fc-structure, which is absent in the nanobodies(6,7). However, as another opportunity, these interesting and stable single-domain antibodies(8) can be easily engineered with different tags. The widely used (His)(6)-tag shows affinity for bivalent ions such as nickel or cobalt, which can on their turn be easily coated on magnetic beads. We therefore developed this simple quantitative affinity capture assay based on cobalt coated magnetic beads. Poliovirus was labeled

  17. A Magnetic Nanoparticle Based Enzyme-Linked Immunosorbent Assay for Sensitive Quantification of Zearalenone in Cereal and Feed Samples

    PubMed Central

    Zhang, Xian; Wang, Xin; Sun, Mengjiao; Zhang, Xiaofeng; Song, Houhui; Yan, Yaxian; Sun, Jianhe; Li, Xiaoliang; Fang, Weihuan

    2015-01-01

    A novel enzyme-linked immunosorbent assay based on magnetic nanoparticles and biotin/streptavidin-HRP (MNP-bsELISA) was developed for rapid and sensitive detection of zearalenone (ZEN). The detection signal was enhanced and the sensitivity of the assay was improved by combined use of antibody-conjugated magnetic nanoparticles and biotin-streptavidin system. Under the optimized conditions, the regression equation for quantification of ZEN was y = −0.4287x + 0.3132 (R2 = 0.9904). The working range was 0.07–2.41 ng/mL. The detection limit was 0.04 ng/mL and IC50 was 0.37 ng/mL. The recovery rates of intra-assay and inter-assay ranged from 92.8%–111.9% and 91.7%–114.5%, respectively, in spiked corn samples. Coefficients of variation were less than 10% in both cases. Parallel analysis of cereal and feed samples showed good correlation between MNP-bsELISA and liquid chromatograph-tandem mass spectrometry (R2 = 0.9283). We conclude that this method is suitable for rapid detection of zearalenone in cereal and feed samples in relevant laboratories. PMID:26492271

  18. Magnetic levitation as a platform for competitive protein-ligand binding assays.

    PubMed

    Shapiro, Nathan D; Soh, Siowling; Mirica, Katherine A; Whitesides, George M

    2012-07-17

    This paper describes a method based on magnetic levitation (MagLev) that is capable of indirectly measuring the binding of unlabeled ligands to unlabeled protein. We demonstrate this method by measuring the affinity of unlabeled bovine carbonic anhydrase (BCA) for a variety of ligands (most of which are benzene sulfonamide derivatives). This method utilizes porous gel beads that are functionalized with a common aryl sulfonamide ligand. The beads are incubated with BCA and allowed to reach an equilibrium state in which the majority of the immobilized ligands are bound to BCA. Since the beads are less dense than the protein, protein binding to the bead increases the overall density of the bead. This change in density can be monitored using MagLev. Transferring the beads to a solution containing no protein creates a situation where net protein efflux from the bead is thermodynamically favorable. The rate at which protein leaves the bead for the solution can be calculated from the rate at which the levitation height of the bead changes. If another small molecule ligand of BCA is dissolved in the solution, the rate of protein efflux is accelerated significantly. This paper develops a reaction-diffusion (RD) model to explain both this observation, and the physical-organic chemistry that underlies it. Using this model, we calculate the dissociation constants of several unlabeled ligands from BCA, using plots of levitation height versus time. Notably, although this method requires no electricity, and only a single piece of inexpensive equipment, it can measure accurately the binding of unlabeled proteins to small molecules over a wide range of dissociation constants (K(d) values within the range from ~10 nM to 100 μM are measured easily). Assays performed using this method generally can be completed within a relatively short time period (20 min-2 h). A deficiency of this system is that it is not, in its present form, applicable to proteins with molecular weight greater

  19. Dual magnetic-/temperature-responsive nanoparticles for microfluidic separations and assays.

    PubMed

    Lai, James J; Hoffman, John M; Ebara, Mitsuhiro; Hoffman, Allan S; Estournès, Claude; Wattiaux, Alain; Stayton, Patrick S

    2007-06-19

    A stimuli-responsive magnetic nanoparticle system for diagnostic target capture and concentration has been developed for microfluidic lab card settings. Telechelic poly(N-isopropylacrylamide) (PNIPAAm) polymer chains were synthesized with dodecyl tails at one end and a reactive carboxylate at the opposite end by the reversible addition fragmentation transfer technique. These PNIPAAm chains self-associate into nanoscale micelles that were used as dimensional confinements to synthesize the magnetic nanoparticles. The resulting superparamagnetic nanoparticles exhibit a gamma-Fe2O3 core ( approximately 5 nm) with a layer of carboxylate-terminated PNIPAAm chains as a corona on the surface. The carboxylate group was used to functionalize the magnetic nanoparticles with biotin and subsequently with streptavidin. The functionalized magnetic nanoparticles can be reversibly aggregated in solution as the temperature is cycled through the PNIPAAm lower critical solution temperature (LCST). While the magnetophoretic mobility of the individual nanoparticles below the LCST is negligible, the aggregates formed above the LCST are large enough to respond to an applied magnetic field. The magnetic nanoparticles can associate with biotinylated targets as individual particles, and then subsequent application of a combined temperature increase and magnetic field can be used to magnetically separate the aggregated particles onto the poly(ethylene glycol)-modified polydimethylsiloxane channel walls of a microfluidic device. When the magnetic field is turned off and the temperature is reversed, the captured aggregates redisperse into the channel flow stream for further downstream processing. The dual magnetic- and temperature-responsive nanoparticles can thus be used as soluble reagents to capture diagnostic targets at a controlled time point and channel position. They can then be isolated and released after the nanoparticles have captured target molecules, overcoming the problem of low

  20. A spheroid toxicity assay using magnetic 3D bioprinting and real-time mobile device-based imaging.

    PubMed

    Tseng, Hubert; Gage, Jacob A; Shen, Tsaiwei; Haisler, William L; Neeley, Shane K; Shiao, Sue; Chen, Jianbo; Desai, Pujan K; Liao, Angela; Hebel, Chris; Raphael, Robert M; Becker, Jeanne L; Souza, Glauco R

    2015-01-01

    An ongoing challenge in biomedical research is the search for simple, yet robust assays using 3D cell cultures for toxicity screening. This study addresses that challenge with a novel spheroid assay, wherein spheroids, formed by magnetic 3D bioprinting, contract immediately as cells rearrange and compact the spheroid in relation to viability and cytoskeletal organization. Thus, spheroid size can be used as a simple metric for toxicity. The goal of this study was to validate spheroid contraction as a cytotoxic endpoint using 3T3 fibroblasts in response to 5 toxic compounds (all-trans retinoic acid, dexamethasone, doxorubicin, 5'-fluorouracil, forskolin), sodium dodecyl sulfate (+control), and penicillin-G (-control). Real-time imaging was performed with a mobile device to increase throughput and efficiency. All compounds but penicillin-G significantly slowed contraction in a dose-dependent manner (Z' = 0.88). Cells in 3D were more resistant to toxicity than cells in 2D, whose toxicity was measured by the MTT assay. Fluorescent staining and gene expression profiling of spheroids confirmed these findings. The results of this study validate spheroid contraction within this assay as an easy, biologically relevant endpoint for high-throughput compound screening in representative 3D environments. PMID:26365200

  1. A spheroid toxicity assay using magnetic 3D bioprinting and real-time mobile device-based imaging

    PubMed Central

    Tseng, Hubert; Gage, Jacob A.; Shen, Tsaiwei; Haisler, William L.; Neeley, Shane K.; Shiao, Sue; Chen, Jianbo; Desai, Pujan K.; Liao, Angela; Hebel, Chris; Raphael, Robert M.; Becker, Jeanne L.; Souza, Glauco R.

    2015-01-01

    An ongoing challenge in biomedical research is the search for simple, yet robust assays using 3D cell cultures for toxicity screening. This study addresses that challenge with a novel spheroid assay, wherein spheroids, formed by magnetic 3D bioprinting, contract immediately as cells rearrange and compact the spheroid in relation to viability and cytoskeletal organization. Thus, spheroid size can be used as a simple metric for toxicity. The goal of this study was to validate spheroid contraction as a cytotoxic endpoint using 3T3 fibroblasts in response to 5 toxic compounds (all-trans retinoic acid, dexamethasone, doxorubicin, 5′-fluorouracil, forskolin), sodium dodecyl sulfate (+control), and penicillin-G (−control). Real-time imaging was performed with a mobile device to increase throughput and efficiency. All compounds but penicillin-G significantly slowed contraction in a dose-dependent manner (Z’ = 0.88). Cells in 3D were more resistant to toxicity than cells in 2D, whose toxicity was measured by the MTT assay. Fluorescent staining and gene expression profiling of spheroids confirmed these findings. The results of this study validate spheroid contraction within this assay as an easy, biologically relevant endpoint for high-throughput compound screening in representative 3D environments. PMID:26365200

  2. Photocleavable Peptide-Conjugated Magnetic Beads for Protein Kinase Assays by MALDI-TOF MS

    PubMed Central

    Zhou, Guangchang; Yan, Xiaoliang; Wu, Ding; Kron, Stephen J.

    2015-01-01

    Peptides were immobilized onto superparamagnetic beads via photocleavable linkers. This enabled simple, rapid, and label-free protein kinase assays via MALDI-TOF MS detection of substrate peptide phosphorylation. Abltide, a model substrate for the Abl protein tyrosine kinase model, was coupled onto amine-terminated beads, incubated with ATP and recombinant c-Abl kinase, and released and further detected to determine phosphorylation. Abltide phosphorylation was found to depend significantly on the length and composition of linkers to the bead surface. Inserting a diblock spacer of poly(glycine) and poly(ethylene glycol) segments markedly enhanced phosphorylation. To validate the assay, the activity of two small-molecule kinase inhibitors, imatinib and dasatinib, which target the oncogenic mutant tyrosine kinase Bcr-Abl to treat chronic myeloid leukemia (CML), was tested. Examining inhibition of the purified c-Abl or Bcr-Abl in K562 CML cell extracts, IC50 values were determined to be consistent with the literature. This simple, label-free, MALDI-based protein kinase assay can be readily adapted to allow multiplexed assays of multiple peptide substrates and/or analysis of alternative post-translational modifications as a tool for drug discovery and clinical testing. PMID:20860375

  3. Dual-readout fluorescent assay of protein kinase activity by use of TiO2-coated magnetic microspheres.

    PubMed

    Bai, Jie; Zhao, Yunjie; Wang, Zhibin; Liu, Chenghui; Wang, Yucong; Li, Zhengping

    2013-05-01

    A simple, highly sensitive, and dual-readout fluorescent assay is developed for the detection of protein kinase activity based on the specific recognition utility of TiO2-coated Fe3O4/SiO2 magnetic microspheres (TMSPs) for kinase-induced phosphopeptides. When the fluorophore-labeled substrate peptides are phosphorylated by the kinase reaction, they can bind specifically to the TiO2 layer of TMSPs by means of phosphate groups, resulting in fluorophore enrichment on the TMSP surfaces. The accumulated fluorophores on the TMSPs are proportional to the kinase activity, and the fluorescence signal readout could be run through either direct fluorescent imaging of the TMSPs or measurement of the fluorescence intensity by simply detaching the fluorescent phosphopeptides into the solution. The TMSPs exhibit extremely high selectivity for capturing phosphorylated peptides over the nonphosphorylated ones, resulting in an ultrahigh fluorescence signal-to-background ratio of 42, which is the highest fluorescence change thus far in fluorescent assays for detection of protein kinase activities. Therefore, the proposed fluorescent assay presents high sensitivity, low detection limit of 0.1 milliunit/μL, and wide dynamic range from 0.5 milliunit/μL to 0.5 unit/μL with protein kinase A (PKA) as a model target. Moreover, the TMSP-based fluorescent assay can simultaneously quantify multiple kinase activities with their specific peptides labeled with different dyes. This new strategy is also successfully applied to monitoring drug-triggered PKA activation in cell lysates. Therefore, the TMSP-based fluorescent assay is very promising in high-throughput screening of kinase inhibitors and in highly sensitive detection of kinase activity, and thus it is a valuable tool for development of targeted therapy, clinical diagnosis, and studies of fundamental life science. PMID:23581884

  4. Magnetically optimized SERS assay for rapid detection of trace drug-related biomarkers in saliva and fingerprints.

    PubMed

    Yang, Tianxi; Guo, Xiaoyu; Wang, Hui; Fu, Shuyue; Wen, Ying; Yang, Haifeng

    2015-06-15

    New developments in the fields of human healthcare and social security call for the exploration of an easy and on-field method to detect drug-related biomarkers. In this paper, Au nanoparticles dotted magnetic nanocomposites (AMN) modified with inositol hexakisphosphate (IP6) were used as surface-enhanced Raman scattering (SERS) substrate to quickly monitor trace drug-related biomarkers in saliva and to on-site screen a trace drug biomarker in fingerprints. Due to inducing with an external magnet, such substrate presented a huge SERS activity, which has met the sensitivity requirement for assay to detect the drug biomarkers in saliva from the U.S. Substance Abuse and Mental Health Services Administration, and also the limit of detection for drug biomarker in fingerprint reached 100 nM. In addition, this AMN-based SERS assay was successfully conducted using a portable Raman spectrometer, which could be used to on-site and accurately differentiate between the smokers and drug addicts in near future.

  5. A novel assay for screening inhibitors targeting HIV-1 integrase dimerization based on Ni-NTA magnetic agarose beads.

    PubMed

    Zhang, Dawei; He, Hongqiu; Liu, Mengmeng; Meng, Zhixia; Guo, Shunxing

    2016-05-03

    Human immunodeficiency virus (HIV)-1 integrase (IN), which mediates integration of viral cDNA into the cellular chromosome, is a validated antiviral drug target. Three IN inhibitors, raltegravir, elvitegravir and dolutegravir, have been clinically approved since 2008. However, drug resistance have emerged in infected patients receiving treatment using these drugs which share the same mechanism of action and have a low genetic barrier for resistance. Therefore, there is an urgent need to develop drugs with novel mechanism. IN requires a precise and dynamic equilibrium between several oligomeric species for its activities. The modulation of the process which is termed as IN oligomerization, presents an interesting allosteric target for drug development. In this research, we developed a magnetic beads based approach to assay the IN dimerization. Then, using the assay we screened a library of 1000 Food and Drug Administration (FDA)-approved drugs for IN dimerization inhibitors and identified dexlansoprazole as a potential IN dimerization inhibitor. In conclusion, the assay presented here has been proven to be sensitive and specific for the detection of IN dimerization as well as for the identification of antiviral drugs targeting IN dimerization. Moreover, a FDA-approved proton-pump inhibitors, dexlansoprazole, was identified as a potential inhibitor for IN dimerization.

  6. Development of a magnetic capture hybridization real-time PCR assay for detection of tumorigenic Agrobacterium vitis in grapevines.

    PubMed

    Johnson, Kameka L; Zheng, Desen; Kaewnum, Supaporn; Reid, Cheryl Lynn; Burr, Thomas

    2013-06-01

    Agrobacterium vitis, the causal agent of grape crown gall, can have severe economic effects on grape production. The bacterium survives systemically in vines and, therefore, is disseminated in propagation material. We developed an assay for use in indexing programs that is efficient and sensitive for detecting A. vitis in grape tissue. Initially, real-time polymerase chain reaction (PCR) primers specific for diverse tumorigenic strains of A. vitis were developed using the virD2 gene sequence. To overcome the effects of PCR inhibitors present in plant tissue, DNA extraction methods that included magnetic capture hybridization (MCH), immunomagnetic separation (IMS), and extraction with the Mo Bio Powerfood kit were compared. The assays incorporating MCH or IMS followed by real-time PCR were 10,000-fold more sensitive than direct real-time PCR when tested using boiled bacterial cell suspensions, with detection thresholds of 10(1) CFU/ml compared with 10(5) CFU/ml. DNA extraction with the Powerfood DNA extraction kit was 10-fold more sensitive than direct real-time PCR, with a detection threshold of 10(4) CFU/ml. All three assays were able to detect A. vitis in healthy-appearing grapevine cuttings taken from infected vines. PMID:23324046

  7. A novel assay for screening inhibitors targeting HIV-1 integrase dimerization based on Ni-NTA magnetic agarose beads

    PubMed Central

    Zhang, Dawei; He, Hongqiu; Liu, Mengmeng; Meng, Zhixia; Guo, Shunxing

    2016-01-01

    Human immunodeficiency virus (HIV)-1 integrase (IN), which mediates integration of viral cDNA into the cellular chromosome, is a validated antiviral drug target. Three IN inhibitors, raltegravir, elvitegravir and dolutegravir, have been clinically approved since 2008. However, drug resistance have emerged in infected patients receiving treatment using these drugs which share the same mechanism of action and have a low genetic barrier for resistance. Therefore, there is an urgent need to develop drugs with novel mechanism. IN requires a precise and dynamic equilibrium between several oligomeric species for its activities. The modulation of the process which is termed as IN oligomerization, presents an interesting allosteric target for drug development. In this research, we developed a magnetic beads based approach to assay the IN dimerization. Then, using the assay we screened a library of 1000 Food and Drug Administration (FDA)-approved drugs for IN dimerization inhibitors and identified dexlansoprazole as a potential IN dimerization inhibitor. In conclusion, the assay presented here has been proven to be sensitive and specific for the detection of IN dimerization as well as for the identification of antiviral drugs targeting IN dimerization. Moreover, a FDA-approved proton-pump inhibitors, dexlansoprazole, was identified as a potential inhibitor for IN dimerization. PMID:27137477

  8. Sensitive electrochemical determination of miRNAs based on a sandwich assay onto magnetic microcarriers and hybridization chain reaction amplification.

    PubMed

    Torrente-Rodríguez, R M; Campuzano, S; Montiel, V Ruiz-Valdepeñas; Montoya, J J; Pingarrón, J M

    2016-12-15

    A novel electrochemical approach for determination of miRNAs involving a sandwich hybridization assay onto streptavidin-magnetic beads (Strep-MBs), hybridization chain reaction (HCR) amplification and amperometric detection at disposable screen-printed carbon electrodes is reported. Using miRNA-21 as the target analyte, a dynamic linear range from 0.2 to 5.0nM with a 60pM (1.5fmol in 25μL) detection limit was obtained. The achieved sensitivity is 24-fold higher than a non-HCR amplification approach involving conventional sandwich type assay onto MBs. Moreover, the whole assay time lasted 1h 45min which is remarkably shorter than other reported methodologies. The methodology exhibited full selectivity against other non-complementary miRNAs as well as an acceptable discrimination between homologous miRNA family members. The applicability of this novel approach was demonstrated by determining mature miRNA-21 in total RNA (RNAt) extracted from tumor cells and human tissues.

  9. Study of the temperature dependent immuno-reaction kinetics for the bio-functionalized magnetic nanoparticle assay of bio-markers of colorectal cancer

    NASA Astrophysics Data System (ADS)

    Yang, S. Y.; Chang, J. F.; Chen, T. C.; Yang, C. C.; Ho, C. S.

    2014-01-01

    By conjugating antibodies on magnetic nanoparticles, target antigens can be quantitatively detected by measuring the magnetic signals of the magnetic nanoparticles due to their association with target antigens. This method of detection is called magnetically labeled immunoassay. The assay characteristics of magnetically labeled immunoassay have been reported widely. However, the immuno-reaction kinetics of magnetically labeled immunoassay has not been studied. In this work, the reaction rates between magnetic nanoparticles and target antigens are measured at various temperatures. It is found that the temperature dependent reaction rate obeys Arrhenius's equation, which shows the collision frequency and activation energy for the immuno-reaction between antibody-functionalized magnetic nanoparticles and target antigens. The carcinoembryonic antigen, which is a regular blood bio-marker for in-vitro diagnosis of colorectal cancer, is used as a target antigen for the example.

  10. Discrimination of clostridium species using a magnetic bead based hybridization assay

    NASA Astrophysics Data System (ADS)

    Pahlow, Susanne; Seise, Barbara; Pollok, Sibyll; Seyboldt, Christian; Weber, Karina; Popp, Jürgen

    2014-05-01

    Clostridium chauvoei is the causative agent of blackleg, which is an endogenous bacterial infection. Mainly cattle and other ruminants are affected. The symptoms of blackleg are very similar to those of malignant edema, an infection caused by Clostridium septicum. [1, 2] Therefore a reliable differentiation of Clostridium chauvoei from other Clostridium species is required. Traditional microbiological detection methods are time consuming and laborious. Additionally, the unique identification is hindered by the overgrowing tendency of swarming Clostridium septicum colonies when both species are present. [1, 3, 4] Thus, there is a crucial need to improve and simplify the specific detection of Clostridium chauvoei and Clostridium septicum. Here we present an easy and fast Clostridium species discrimination method combining magnetic beads and fluorescence spectroscopy. Functionalized magnetic particles exhibit plentiful advantages, like their simple manipulation in combination with a large binding capacity of biomolecules. A specific region of the pathogenic DNA is amplified and labelled with biotin by polymerase chain reaction (PCR). These PCR products were then immobilized on magnetic beads exploiting the strong biotin-streptavidin interaction. The specific detection of different Clostridium species is achieved by using fluorescence dye labeled probe DNA for the hybridization with the immobilized PCR products. Finally, the samples were investigated by fluorescence spectroscopy. [5

  11. Colorimetric Assay for the Detection of Typical Biomarkers for Periodontitis Using a Magnetic Nanoparticle Biosensor.

    PubMed

    Wignarajah, Shayalini; Suaifan, Ghadeer A R Y; Bizzarro, Sergio; Bikker, Floris J; Kaman, Wendy E; Zourob, Mohammed

    2015-12-15

    Periodontitis is a chronic disease which affects at least 10% of the population. If untreated, periodontitis can lead to teeth loss. Unfortunately, current diagnostic tests are limited in their sensitivity and specificity. In this study, a novel multiplex hand-held colorimetric diagnostic biosensor, using two typical inflammatory salivary biomarkers, Human Neutrophil Elastase (HNE) and Cathepsin-G, was constructed as proof of concept to potentially detect periodontitis. The biosensing method was based on the measurement of proteolytic activity using specific proteases probes. These probes consist of specific proteases substrates covalently bound to a magnetic bead from one end and to the gold sensor surface by the other end. When intact, this renders the golden sensor black. Upon proteolysis, the cleaved magnetic beads will be attracted by an external magnet revealing the golden color of the sensor surface observable by the naked eye. The biosensor was capable of specific and quantitative detection of HNE and Cathepsin-G in solution and in spiked saliva samples with a lower detection limit of 1 pg/mL and 100 fg/mL for HNE and Cathepsin-G, respectively. Examination of periodontitis patients' sample and a healthy control showed the potential of the multiplex biosensor to detect the presence of HNE and Cathepsin-G activity in situ. This approach is anticipated to be a useful biochip array amenable to low-cost point-of-care devices.

  12. Development of SERS substrate using phage-based magnetic template for triplex assay in sepsis diagnosis.

    PubMed

    Nguyen, Anh H; Shin, Yesol; Sim, Sang Jun

    2016-11-15

    Development of a new substrate for surface-enhanced Raman scattering (SERS) is one area of interest for the improvement of SERS performance. Herein, we introduce a new method for developing new mesoporous SERS substrates using M13 phages that display cysteine-rich peptides on the pVIII major units, which is an alternative for thiol donor using chemical modifications. Together with the SERS substrate development, and the use of the SERS technique for sepsis diagnostics is a new approach in clinical settings. The substrates were characterized and magnetized with magnetic immuno colloids made of gold-coated magnetic nanoparticles and specific antibodies. Conventionally, the SERS-tags are prepared by using gold nanoparticles and are modified with Raman dyes to immobilize specific antibodies to capture the biomarkers in the serum samples. However, in this method the SERS-tags are bound to the mesoporous substrate via antibody/antigen interactions to form clusters or layer-by-layer assemblies of SERS-tags for Raman signal enhancement. The SERS spectra showed distinct peaks for tags corresponding to three typical sepsis-specific biomarkers for diagnostics with the limit of detection values of 27 pM, 103 pM, and 78 pM for C-reactive protein (CRP), procalcitonin (PCT), and soluble triggering receptor expressed on myeloid cells-1 (sTREM-1), respectively. With such an approach, SERS can be used for clinical purposes and can be improved by phage display modification rather than chemical alternatives. PMID:27209579

  13. Magnetic-nanobead-based competitive enzyme-linked aptamer assay for the analysis of oxytetracycline in food.

    PubMed

    Lu, Chunxia; Tang, Zonggui; Liu, Changbin; Kang, Lichao; Sun, Fengxia

    2015-05-01

    This study presents a novel analytical method for the detection of oxytetracycline (OTC) in complex food matrices based on a direct competitive enzyme-linked aptamer assay and magnetic separation technology. In this protocol, free OTC competed with horseradish peroxidase labeled OTC (OTC-HRP) for binding to the OTC aptamer immobilized on magnetic beads. The parameters that can affect the response, such as avidin concentration, aptamer concentration, OTC-HRP concentration, incubation temperature, incubation time, blocking agent, and binding buffer, were optimized. Under the optimal conditions, the linear range for the OTC concentration detection is 0.5-100 ng mL(-1), with a concentration of OTC needed to obtain 50 % of the maximum signal of 14.47 ng mL(-1). The limit of detection and the limit of quantitation were 0.88 and 3.40 ng mL(-1), respectively. There was no obvious cross-reactivity with most of the tetracycline pesticides. The recovery rates ranged from 71.0 to 91.2 % for the food samples, including chicken, milk, and honey, and the relative standard deviation was less than 15.0 %. The proposed method was applied to measure OTC in real samples, and was validated using high-performance liquid chromatography. This method has the advantages of magnetic separation and the concentration effect of magnetic nanoparticles, the specificity of the aptamer, and the high-throughput of microtiter plates; it offers a promising approach for the screening of OTC because it is simple, rapid, highly sensitive, and has low cost.

  14. Magnetic-nanobead-based competitive enzyme-linked aptamer assay for the analysis of oxytetracycline in food.

    PubMed

    Lu, Chunxia; Tang, Zonggui; Liu, Changbin; Kang, Lichao; Sun, Fengxia

    2015-05-01

    This study presents a novel analytical method for the detection of oxytetracycline (OTC) in complex food matrices based on a direct competitive enzyme-linked aptamer assay and magnetic separation technology. In this protocol, free OTC competed with horseradish peroxidase labeled OTC (OTC-HRP) for binding to the OTC aptamer immobilized on magnetic beads. The parameters that can affect the response, such as avidin concentration, aptamer concentration, OTC-HRP concentration, incubation temperature, incubation time, blocking agent, and binding buffer, were optimized. Under the optimal conditions, the linear range for the OTC concentration detection is 0.5-100 ng mL(-1), with a concentration of OTC needed to obtain 50 % of the maximum signal of 14.47 ng mL(-1). The limit of detection and the limit of quantitation were 0.88 and 3.40 ng mL(-1), respectively. There was no obvious cross-reactivity with most of the tetracycline pesticides. The recovery rates ranged from 71.0 to 91.2 % for the food samples, including chicken, milk, and honey, and the relative standard deviation was less than 15.0 %. The proposed method was applied to measure OTC in real samples, and was validated using high-performance liquid chromatography. This method has the advantages of magnetic separation and the concentration effect of magnetic nanoparticles, the specificity of the aptamer, and the high-throughput of microtiter plates; it offers a promising approach for the screening of OTC because it is simple, rapid, highly sensitive, and has low cost. PMID:25855149

  15. Thrombin-linked aptamer assay for detection of platelet derived growth factor BB on magnetic beads in a sandwich format.

    PubMed

    Guo, Limin; Zhao, Qiang

    2016-09-01

    Here we describe a thrombin-linked aptamer assay (TLAA) for protein by using thrombin as an enzyme label, harnessing enzyme activity of thrombin and aptamer affinity binding. TLAA converts detection of specific target proteins to the detection of thrombin by using a DNA sequence that consists of two aptamers with the first aptamer binding to the specific target protein and the second aptamer binding to thrombin. Through the affinity binding, the thrombin enzyme is labeled on the protein target, and thrombin catalyzes the hydrolysis of small peptide substrate into product, generating signals for quantification. As a proof of principle, we show a sandwich TLAA for platelet derived growth factor BB (PDGF-BB) by using anti-PDGF-BB antibody coated on magnetic beads and an oligonucleotide containing the aptamer for PDGF-BB and the aptamer for thrombin. The binding of PDGF-BB to both the antibody and the aptamer results in labeling the complex with thrombin. We achieved detection of PDGF-BB at 16 pM. This TLAA contributes a new application of thrombin and its aptamer in bioanalysis, and shows potentials in assay developments. PMID:27343590

  16. One-step multiplexed detection of foodborne pathogens: Combining a quantum dot-mediated reverse assaying strategy and magnetic separation.

    PubMed

    Yin, Binfeng; Wang, Yu; Dong, Mingling; Wu, Jing; Ran, Bei; Xie, Mengxia; Joo, Sang Woo; Chen, Yiping

    2016-12-15

    A rapid and multiplexed immunosensor was developed based on a quantum dot (QD)-reverse assaying strategy (RAS) and immuno-magnetic beads (IMBs) for one-step and simultaneous detection of Escherichia coli O157: H7 and Salmonella. In a conventional QD-based immunosensor, the fluorescence signal of the "IMBs-target-QD" immunoconjugate is directly used as the assaying readout. However, the fluorescence signal is affected by IMBs due to light scattering and the "IMBs-target-QD" immunoconjugate needs multiple washing and re-suspension steps. To address these problems, we use the surplus QD-antibody conjugate as signal readout in the RAS, which prevents interference from the IMBs, increases the fluorescence signal, and avoids complex operations. Compared with conventional QD-based immunosensor, the sensitivity of QD-RSA immunosensor for detection of Escherichia coli O157: H7 has been improved fifty-fold, and whole analysis procedure can be finished within 1h. Therefore, this RSA strategy is promising for improving the performance of QD-based immunosensors and could greatly broaden their applications.

  17. An aptamer-based assay for thrombin via structure switch based on gold nanoparticles and magnetic nanoparticles.

    PubMed

    Zheng, Jing; Cheng, Gui-Fang; He, Pin-Gang; Fang, Yu-Zhi

    2010-03-15

    An aptamer-based assay for thrombin with high specificity and sensitivity was presented. In the protocol, the aptamer for thrombin was immobilized on magnetic nanoparticle, and its complementary oligonucleotide was labeled with gold nanoparticles, then the aptamer was hybridized with the complementary oligonucleotide to form the duplex structure as a probe, this probe could be used for the specific recognition for thrombin. In the presence of thrombin, the aptamer prefer to form the G-quarter structure with thrombin, resulting in the dissociation of the duplex of the probe and the release of the gold labeled oligonucleotide. Upon this, we were able to detect thrombin through the detection of the electrochemical signal of gold nanoparticles. The strategy combines with the high specificity of aptamer and the excellent characteristics of nanoparticles. This assay is simple, rapid, sensitive and highly specific, it does not require labeling of thrombin, and it could be applied to detect thrombin in complex real sample. The method shows great potential in other protein analysis and in disease diagnosis.

  18. One-step multiplexed detection of foodborne pathogens: Combining a quantum dot-mediated reverse assaying strategy and magnetic separation.

    PubMed

    Yin, Binfeng; Wang, Yu; Dong, Mingling; Wu, Jing; Ran, Bei; Xie, Mengxia; Joo, Sang Woo; Chen, Yiping

    2016-12-15

    A rapid and multiplexed immunosensor was developed based on a quantum dot (QD)-reverse assaying strategy (RAS) and immuno-magnetic beads (IMBs) for one-step and simultaneous detection of Escherichia coli O157: H7 and Salmonella. In a conventional QD-based immunosensor, the fluorescence signal of the "IMBs-target-QD" immunoconjugate is directly used as the assaying readout. However, the fluorescence signal is affected by IMBs due to light scattering and the "IMBs-target-QD" immunoconjugate needs multiple washing and re-suspension steps. To address these problems, we use the surplus QD-antibody conjugate as signal readout in the RAS, which prevents interference from the IMBs, increases the fluorescence signal, and avoids complex operations. Compared with conventional QD-based immunosensor, the sensitivity of QD-RSA immunosensor for detection of Escherichia coli O157: H7 has been improved fifty-fold, and whole analysis procedure can be finished within 1h. Therefore, this RSA strategy is promising for improving the performance of QD-based immunosensors and could greatly broaden their applications. PMID:27498327

  19. Magnetic beads-based chemiluminescent assay for ultrasensitive detection of pseudorabies virus.

    PubMed

    Yang, Haowen; Guo, Yafen; Li, Song; Lan, Ganqiu; Jiang, Qinyang; Yang, Xiurong; Fan, Jing; Ali, Zeeshan; Tang, Yongjun; Mou, Xianbo; Liu, Hongna; Shah, Muhammad Ali A; Jin, Si; Jiang, Hesheng; Li, Zhiyang

    2014-05-01

    A rapid, ultrasensitive and economical Pseudorabies virus (PRV) detection system based on magnetic beads (MBs) and chemiluminescence was developed in this paper. The carboxyl functionalized MBs (MBs-COOH) were covalently coupled with aminated DNA probes for capturing PRV biotinylated amplicon, the product of polymerase chain reaction (PCR). Agarose gel electrophoresis analysis approved the reliability of biotinylated amplicon. The MBs composites were incubated with alkaline phosphatase labeled streptavidin (ALP-SA) and chemiluminescene was determined by subsequently adding 3-(2'-spiroadamantane)-4-methoxy-4-(3"-phosphoryloxy)phenyl-1,2-dioxetane (AMPPD). The optimal conditions of the PRV detection method were 10 microM for probe concentration, 50 degrees C for hybridization temperature and 30 min for hybridization time. The limit of detection (LOD) was as low as 100 amol/5 pM of amplicon which proved that this approach for PRV detection was ultrasensitive.

  20. A new gravity-driven microfluidic-based electrochemical assay coupled to magnetic beads for nucleic acid detection.

    PubMed

    Laschi, Serena; Miranda-Castro, Rebeca; González-Fernández, Eva; Palchetti, Ilaria; Reymond, Frédéric; Rossier, Joël S; Marrazza, Giovanna

    2010-11-01

    In this work, the characterisation and the optimisation of hybridisation assays based on a novel, rapid and sensitive micro-analytical, gravity-driven, flow device is reported. This device combines a special chip containing eight polymer microchannels, with a portable, computer-controlled instrument. The device is used as a platform for affinity experiments using oligonucleotide-modified paramagnetic particles. In our approach, both hybridisation and labelling events are performed on streptavidin-coated paramagnetic microparticles functionalized with a biotinylated capture probe. Modified particles, introduced in the microchannel inlet of the chip, accumulate near the electrode surface by virtue of a magnetic holder. After hybridisation with the complementary sequence, the hybrid is labelled with an alkaline phosphatase conjugate. The electrochemical substrate for alkaline phosphatase revelation is p-aminophenyl phosphate. Solutions and reagents are sequentially passed through the microchannels, until enzyme substrate is added for in situ signal detection. Upon readout, the magnet array is flipped away, beads are removed by addition of regeneration buffer, and the so-regenerated chip is ready for further analysis. This protocol has been applied to the analytical detection of specific DNA sequences of Legionella pneumophila, with an RSD=8.5% and a detection limit of 0.33 nM.

  1. Development of a fluorescent enzyme-linked DNA aptamer-magnetic bead sandwich assay and portable fluorometer for sensitive and rapid leishmania detection in sandflies.

    PubMed

    Bruno, John G; Richarte, Alicia M; Phillips, Taylor; Savage, Alissa A; Sivils, Jeffrey C; Greis, Alex; Mayo, Michael W

    2014-01-01

    A fluorescent peroxidase-linked DNA aptamer-magnetic bead sandwich assay is described which detects as little as 100 ng of soluble protein extracted from Leishmania major promastigotes with a high molarity chaotropic salt. Lessons learned during development of the assay are described and elucidate the pros and cons of using fluorescent dyes or nanoparticles and quantum dots versus a more consistent peroxidase-linked Amplex Ultra Red (AUR; similar to resazurin) fluorescence version of the assay. While all versions of the assays were highly sensitive, the AUR-based version exhibited lower variability between tests. We hypothesize that the AUR version of this assay is more consistent, especially at low analyte levels, because the fluorescent product of AUR is liberated into bulk solution and readily detectable while fluorophores attached to the reporter aptamer might occasionally be hidden behind magnetic beads near the detection limit. Conversely, fluorophores could be quenched by nearby beads or other proximal fluorophores on the high end of analyte concentration, if packed into a small area after magnetic collection when an enzyme-linked system is not used. A highly portable and rechargeable battery-operated fluorometer with on board computer and color touchscreen is also described which can be used for rapid (<1 h) and sensitive detection of Leishmania promastigote protein extracts (∼ 100 ng per sample) in buffer or sandfly homogenates for mapping of L. major parasite geographic distributions in wild sandfly populations.

  2. Development of a fluorescent enzyme-linked DNA aptamer-magnetic bead sandwich assay and portable fluorometer for sensitive and rapid listeria detection.

    PubMed

    Bruno, John G; Phillips, Taylor; Montez, Tiffany; Garcia, Adrian; Sivils, Jeffrey C; Mayo, Michael W; Greis, Alex

    2015-01-01

    A fluorescent DNA aptamer-magnetic bead sandwich assay was developed to detect listeriolysin O (LLO) protein from pathogenic Listeria bacteria using a peroxidase-linked system, Amplex Ultra Red (AUR; derivatized resazurin) substrate, and a custom-designed handheld fluorometer. The assay is highly sensitive with demonstrated limits of detection (LODs) in the range of 4 to 61 L. monocytogenes cells or the equivalent LLO produced by 4 to 61 cells on average in separate titration trials. Total assay processing and analysis time was approximately 30 mins. The assay has demonstrated the ability to detect 6 species of Listeria as desired by the USDA's Food Safety Inspection Service (FSIS). The portable system was designed to be used primarily with surface swab samples from fomites, but it can also be used to assess enrichment cultures. The minimal time to detect a positive enrichment culture in our hands from an initial 10 cell inoculum in 200 ml of broth has been 8 h post-incubation at 37 °C in shaker flask cultures. An optional automated magnetic bead assay processing and wash device capable of simultaneously processing 6 samples with low and consistent fluorescence background for higher volume central laboratories is also described.

  3. Development of a fluorescent enzyme-linked DNA aptamer-magnetic bead sandwich assay and portable fluorometer for sensitive and rapid listeria detection.

    PubMed

    Bruno, John G; Phillips, Taylor; Montez, Tiffany; Garcia, Adrian; Sivils, Jeffrey C; Mayo, Michael W; Greis, Alex

    2015-01-01

    A fluorescent DNA aptamer-magnetic bead sandwich assay was developed to detect listeriolysin O (LLO) protein from pathogenic Listeria bacteria using a peroxidase-linked system, Amplex Ultra Red (AUR; derivatized resazurin) substrate, and a custom-designed handheld fluorometer. The assay is highly sensitive with demonstrated limits of detection (LODs) in the range of 4 to 61 L. monocytogenes cells or the equivalent LLO produced by 4 to 61 cells on average in separate titration trials. Total assay processing and analysis time was approximately 30 mins. The assay has demonstrated the ability to detect 6 species of Listeria as desired by the USDA's Food Safety Inspection Service (FSIS). The portable system was designed to be used primarily with surface swab samples from fomites, but it can also be used to assess enrichment cultures. The minimal time to detect a positive enrichment culture in our hands from an initial 10 cell inoculum in 200 ml of broth has been 8 h post-incubation at 37 °C in shaker flask cultures. An optional automated magnetic bead assay processing and wash device capable of simultaneously processing 6 samples with low and consistent fluorescence background for higher volume central laboratories is also described. PMID:25511112

  4. Development of a monoclonal antibody against deoxynivalenol for magnetic nanoparticle-based extraction and an enzyme-linked immunosorbent assay.

    PubMed

    Lee, Hyuk-Mi; Song, Sung-Ok; Cha, Sang-Ho; Wee, Sung-Bok; Bischoff, Karyn; Park, Sung-Won; Son, Seong-Wan; Kang, Hwan-Goo; Cho, Myung-Haing

    2013-01-01

    Monoclonal antibody (mAb, NVRQS-DON) against deoxynivalenol (DON) was prepared. DON-Ag coated enzyme linked immunosorbent assay (ELISA) and DON-Ab coated ELISA were prepared by coating the DON-BSA and DON mAb. Quantitative DON calculation ranged from 50 to 4,000 ng/mL for DON-Ab coated ELISA and from 25 to 500 ng/mL for DON-Ag coated ELISA. 50% of inhibitory concentration values of DON, HT-2, 15-acetyl-DON, and nivalenol were 23.44, 22,545, 5,518 and 5,976 ng/mL based on the DON-Ab coated ELISA. Cross-reactivity levels of the mAb to HT-2, 15-acetyl-DON, and nivalenol were 0.1, 0.42, and 0.40%. The intra- and interassay precision coefficient variation (CV) were both <10%. In the mAb-coated ELISA, mean DON recovery rates in animal feed (0 to 1,000 mg/kg) ranged from 68.34 to 95.49% (CV; 4.10 to 13.38%). DON in a buffer solution (250, 500 and 1,000 ng/mL) was isolated using 300 mg of NVRQS-DON and 3 mg of magnetic nanoparticles (MNPs). The mean recovery rates of DON using this mAb-MNP system were 75.2, 96.9, and 88.1% in a buffer solution spiked with DON (250, 500, and 1,000 ng/mL). Conclusively we developed competitive ELISAs for detecting DON in animal feed and created a new tool for DON extraction using mAb-coupled MNPs.

  5. Lab-on-a-disc agglutination assay for protein detection by optomagnetic readout and optical imaging using nano- and micro-sized magnetic beads.

    PubMed

    Uddin, Rokon; Burger, Robert; Donolato, Marco; Fock, Jeppe; Creagh, Michael; Hansen, Mikkel Fougt; Boisen, Anja

    2016-11-15

    We present a biosensing platform for the detection of proteins based on agglutination of aptamer coated magnetic nano- or microbeads. The assay, from sample to answer, is integrated on an automated, low-cost microfluidic disc platform. This ensures fast and reliable results due to a minimum of manual steps involved. The detection of the target protein was achieved in two ways: (1) optomagnetic readout using magnetic nanobeads (MNBs); (2) optical imaging using magnetic microbeads (MMBs). The optomagnetic readout of agglutination is based on optical measurement of the dynamics of MNB aggregates whereas the imaging method is based on direct visualization and quantification of the average size of MMB aggregates. By enhancing magnetic particle agglutination via application of strong magnetic field pulses, we obtained identical limits of detection of 25pM with the same sample-to-answer time (15min 30s) using the two differently sized beads for the two detection methods. In both cases a sample volume of only 10µl is required. The demonstrated automation, low sample-to-answer time and portability of both detection instruments as well as integration of the assay on a low-cost disc are important steps for the implementation of these as portable tools in an out-of-lab setting.

  6. Lab-on-a-disc agglutination assay for protein detection by optomagnetic readout and optical imaging using nano- and micro-sized magnetic beads.

    PubMed

    Uddin, Rokon; Burger, Robert; Donolato, Marco; Fock, Jeppe; Creagh, Michael; Hansen, Mikkel Fougt; Boisen, Anja

    2016-11-15

    We present a biosensing platform for the detection of proteins based on agglutination of aptamer coated magnetic nano- or microbeads. The assay, from sample to answer, is integrated on an automated, low-cost microfluidic disc platform. This ensures fast and reliable results due to a minimum of manual steps involved. The detection of the target protein was achieved in two ways: (1) optomagnetic readout using magnetic nanobeads (MNBs); (2) optical imaging using magnetic microbeads (MMBs). The optomagnetic readout of agglutination is based on optical measurement of the dynamics of MNB aggregates whereas the imaging method is based on direct visualization and quantification of the average size of MMB aggregates. By enhancing magnetic particle agglutination via application of strong magnetic field pulses, we obtained identical limits of detection of 25pM with the same sample-to-answer time (15min 30s) using the two differently sized beads for the two detection methods. In both cases a sample volume of only 10µl is required. The demonstrated automation, low sample-to-answer time and portability of both detection instruments as well as integration of the assay on a low-cost disc are important steps for the implementation of these as portable tools in an out-of-lab setting. PMID:27183287

  7. Magnetic bead-based phage anti-immunocomplex assay (PHAIA) for the detection of the urinary biomarker 3-phenoxybenzoic acid to assess human exposure to pyrethroid insecticides.

    PubMed

    Kim, Hee-Joo; Ahn, Ki Chang; González-Techera, Andrés; González-Sapienza, Gualberto G; Gee, Shirley J; Hammock, Bruce D

    2009-03-01

    Noncompetitive immunoassays are advantageous over competitive assays for the detection of small molecular weight compounds. We recently demonstrated that phage peptide libraries can be an excellent source of immunoreagents that facilitate the development of sandwich-type noncompetitive immunoassays for the detection of small analytes, avoiding the technical challenges of producing anti-immunocomplex antibody. In this work we explore a new format that may help to optimize the performance of the phage anti-immunocomplex assay (PHAIA) technology. As a model system we used a polyclonal antibody to 3-phenoxybenzoic acid (3-PBA) and an anti-immunocomplex phage clone bearing the cyclic peptide CFNGKDWLYC. The assay setup with the biotinylated antibody immobilized onto streptavidin-coated magnetic beads significantly reduced the amount of coating antibody giving identical sensitivity (50% saturation of the signal (SC(50))=0.2-0.4ng/ml) to the best result obtained with direct coating of the antibody on ELISA plates. The bead-based assay tolerated up to 10 and 5% of methanol and urine matrix, respectively. This assay system accurately determined the level of spiked 3-PBA in different urine samples prepared by direct dilution or clean-up with solid-phase extraction after acidic hydrolysis with overall recovery of 80-120%.

  8. Magnetic bead-based phage anti-immunocomplex assay (PHAIA) for the detection of the urinary biomarker 3-phenoxybenzoic acid to assess human exposure to pyrethroid insecticides

    PubMed Central

    Kim, Hee-Joo; Ahn, Ki Chang; González-Techera, Andrés; González-Sapienza, Gualberto G.; Gee, Shirley J.; Hammock, Bruce D.

    2010-01-01

    Noncompetitive immunoassays are advantageous over competitive assays for the detection of small molecular weight compounds. We recently demonstrated that phage peptide libraries can be an excellent source of immunoreagents that facilitate the development of sandwich-type noncompetitive immunoassays for the detection of small analytes, avoiding the technical challenges of producing anti-immunocomplex antibody. In this work we explore a new format that may help to optimize the performance of the phage anti-immunocomplex assay (PHAIA) technology. As a model system we used a polyclonal antibody to 3-phenoxybenzoic acid (3-PBA) and an anti-immunocomplex phage clone bearing the cyclic peptide CFNGKDWLYC. The assay setup with the biotinylated antibody immobilized onto streptavidin-coated magnetic beads significantly reduced the amount of coating antibody giving identical sensitivity (50% saturation of the signal (SC50) = 0.2–0.4 ng/ml) to the best result obtained with direct coating of the antibody on ELISA plates. The bead-based assay tolerated up to 10 and 5% of methanol and urine matrix, respectively. This assay system accurately determined the level of spiked 3-PBA in different urine samples prepared by direct dilution or clean-up with solid-phase extraction after acidic hydrolysis with overall recovery of 80–120%. PMID:19101498

  9. Gold magnetic nanoparticle conjugate-based lateral flow assay for the detection of IgM class antibodies related to TORCH infections.

    PubMed

    Li, Xingxing; Zhang, Qinlu; Hou, Peng; Chen, Mingwei; Hui, Wenli; Vermorken, Alphons; Luo, Zhiyi; Li, Hong; Li, Qin; Cui, Yali

    2015-11-01

    In this study, a lateral flow immunochromatographic assay (LFIA) system for the detection of immunoglobulin M (IgM) antibodies, related to TORCH [(T)oxoplasmosis, (O)ther agents, (R)ubella (also known as German Measles), (C)ytomegalovirus, and (H)erpes simplex virus infections], based on gold magnetic nanoparticles, was established. Following modification with poly(methacrylic acid), the gold magnetic nanoparticles conjugated with an anti‑human IgM antibody (μ‑chain specific) to construct a probe. A lateral flow assay device was constructed based on these conjugates. IgM antibodies to four types of pathogens, notably toxoplasmosis, rubella virus, cytomegalovirus and herpes simplex virus type 2, were detected using this device. Compared with commercial colloidal gold‑based LFIA strips, our method exhibited higher sensitivity. No interference with triglycerides, hemoglobin and bilirubin occurred, and no cross‑reactivity was noted among the four pathogens. The gold magnetic nanoparticle‑LFIA strips were used to assess 41 seropositive and 121 seronegative serum samples. The sensitivity was 100% (162/162) and the specificity was 100% (162/162). This method cannot only be used for the detection of TORCH IgM-specific antibodies, but it can potentially be developed for use in the diagnosis of other acute or recently identified autoimmune diseases. PMID:26329478

  10. Development of a single-antigen magnetic bead assay (SAMBA) for the sensitive detection of HPA-1a alloantibodies using tag-engineered recombinant soluble β3 integrin.

    PubMed

    Skaik, Younis; Battermann, Anja; Hiller, Oliver; Meyer, Oliver; Figueiredo, Constanca; Salama, Abdulgabar; Blasczyk, Rainer

    2013-05-31

    Timely and accurate testing for human platelet antigen 1a (HPA-1a) alloantibodies is vital for clinical diagnosis of neonatal alloimmune thrombocytopenia (NAIT). Current antigen-specific assays used for the detection of HPA-1 alloantibodies are technically very complex and cumbersome for most diagnostic laboratories. Hence, we designed and applied recombinant soluble (rs) β3 integrins displaying HPA-1a or HPA-1b epitopes for the development of a single-antigen magnetic bead assay (SAMBA). Soluble HPA-1a and HPA-1b were produced recombinantly in human embryonic kidney 293 (HEK293) cells and differentially tagged. The recombinant soluble proteins were then immobilized onto paramagnetic beads and used for analysis of HPA-1 alloantibodies by enzyme-linked immunosorbent assay (ELISA). HPA-1a serum samples (n=7) from NAIT patients, inert sera and sera containing non-HPA-1a antibodies were used to evaluate the sensitivity and specificity of the SAMBA. Fusion of V5-His or GS-SBP-His tags to the rsβ3 integrins resulted in high-yield expression. SAMBA was able to detect all HPA-1a and -1b alloantibodies recognized by monoclonal antibody-specific immobilization of platelet antigens assay (MAIPA). No cross-reactions between the sera were observed. Two out of seven of the HPA-1a alloantibody-containing sera demonstrated weak to moderate reactivity in MAIPA but strong signals in SAMBA. SAMBA provides a very reliable method for the detection of HPA-1 antibodies with high specificity and sensitivity. This simple and rapid assay can be adapted for use in any routine laboratory and can be potentially adapted for use on automated systems. PMID:23454035

  11. A Novel Assay for Screening Inhibitors Targeting HIV Integrase LEDGF/p75 Interaction Based on Ni2+ Coated Magnetic Agarose Beads

    PubMed Central

    Dawei, Zhang; Hongqiu, He; Mengmeng, Liu; Zhixia, Meng; Shunxing, Guo

    2016-01-01

    HIV-1 integrase (IN) plays an essential role in viral replication and thus serves as an important target for chemotherapeutic intervention against HIV-1 infection. However, the current three clinical IN inhibitors, raltegravir, elvitegravir and dolutegravir share the same inhibitory mechanism, resulting in a common clinical resistance profile which have emerged in infected patients receiving treatment. Therefore, it is important to develop small molecule inhibitors that impair IN function with distinct mechanisms of action. In this work, a magnetic-beads based biochemical assay targeting the protein-protein interaction (PPI) between HIV IN and the cellular cofactor LEDGF/p75 was developed for identification of HIV-1 IN inhibitors. Furthermore, a library containing 1000 US. Food and Drug Administration (FDA)-approved drugs currently used for human medication was screened to identify inhibitors targeting the PPI. The assay was proved to be quite robust and with the novel assay we successfully identified dexlansoprazole (IC50 of 4.8 μM), a FDA-approved proton pump inhibitor, as a potential inhibitor for the PPI between IN and LEDGF/p75, which bound to the LEDGF/p75 partner with a kinetic dissociation (Kd) constant of 330 nM ± 2.6 nM. PMID:27633629

  12. Quantification and viability assays of Toxoplasma gondii in commercial "Serrano" ham samples using magnetic capture real-time qPCR and bioassay techniques.

    PubMed

    Gomez-Samblas, M; Vílchez, S; Racero, J C; Fuentes, M V; Osuna, A

    2015-04-01

    "Serrano" ham is a typical pork product from the Mediterranean area, highly valued for its flavour. To make Serrano ham, pork undergoes a salting and a subsequent fermentation process known as curing. Certain pigs used for meat production are an important source of Toxoplasma gondii infection in humans. We have developed a method for quantifying and assaying the viability of the T. gondii present in commercial Serrano ham samples. A magnetic capture method for the isolation of T. gondii DNA and a qRT-PCR were used to estimate the T. gondii burden in 475 commercial samples of "Serrano" ham in two presentation formats: ham pieces and sliced ham. The infectivity capacity of T. gondii in positive samples was assayed in mice. The global prevalence of T. gondii was 8.84%, ranging from 32.35% in one of the companies to 0% prevalence in three other companies. The infectivity assays revealed that only 4.84% of the positive samples were infective. To the best of our knowledge this is the first report focussing on the prevalence of T. gondii in commercial "Serrano" ham. The method described here could be useful for producers to guarantee the safety of their products.

  13. A Novel Assay for Screening Inhibitors Targeting HIV Integrase LEDGF/p75 Interaction Based on Ni(2+) Coated Magnetic Agarose Beads.

    PubMed

    Dawei, Zhang; Hongqiu, He; Mengmeng, Liu; Zhixia, Meng; Shunxing, Guo

    2016-01-01

    HIV-1 integrase (IN) plays an essential role in viral replication and thus serves as an important target for chemotherapeutic intervention against HIV-1 infection. However, the current three clinical IN inhibitors, raltegravir, elvitegravir and dolutegravir share the same inhibitory mechanism, resulting in a common clinical resistance profile which have emerged in infected patients receiving treatment. Therefore, it is important to develop small molecule inhibitors that impair IN function with distinct mechanisms of action. In this work, a magnetic-beads based biochemical assay targeting the protein-protein interaction (PPI) between HIV IN and the cellular cofactor LEDGF/p75 was developed for identification of HIV-1 IN inhibitors. Furthermore, a library containing 1000 US. Food and Drug Administration (FDA)-approved drugs currently used for human medication was screened to identify inhibitors targeting the PPI. The assay was proved to be quite robust and with the novel assay we successfully identified dexlansoprazole (IC50 of 4.8 μM), a FDA-approved proton pump inhibitor, as a potential inhibitor for the PPI between IN and LEDGF/p75, which bound to the LEDGF/p75 partner with a kinetic dissociation (Kd) constant of 330 nM ± 2.6 nM. PMID:27633629

  14. A Novel Assay for Screening Inhibitors Targeting HIV Integrase LEDGF/p75 Interaction Based on Ni(2+) Coated Magnetic Agarose Beads.

    PubMed

    Dawei, Zhang; Hongqiu, He; Mengmeng, Liu; Zhixia, Meng; Shunxing, Guo

    2016-09-16

    HIV-1 integrase (IN) plays an essential role in viral replication and thus serves as an important target for chemotherapeutic intervention against HIV-1 infection. However, the current three clinical IN inhibitors, raltegravir, elvitegravir and dolutegravir share the same inhibitory mechanism, resulting in a common clinical resistance profile which have emerged in infected patients receiving treatment. Therefore, it is important to develop small molecule inhibitors that impair IN function with distinct mechanisms of action. In this work, a magnetic-beads based biochemical assay targeting the protein-protein interaction (PPI) between HIV IN and the cellular cofactor LEDGF/p75 was developed for identification of HIV-1 IN inhibitors. Furthermore, a library containing 1000 US. Food and Drug Administration (FDA)-approved drugs currently used for human medication was screened to identify inhibitors targeting the PPI. The assay was proved to be quite robust and with the novel assay we successfully identified dexlansoprazole (IC50 of 4.8 μM), a FDA-approved proton pump inhibitor, as a potential inhibitor for the PPI between IN and LEDGF/p75, which bound to the LEDGF/p75 partner with a kinetic dissociation (Kd) constant of 330 nM ± 2.6 nM.

  15. Investigation of magnetic nanoparticles for the rapid extraction and assay of alpha-emitting radionuclides from urine: demonstration of a novel radiobioassay method.

    PubMed

    O'Hara, Matthew J; Carter, Jennifer C; MacLellan, Jay A; Warner, Cynthia L; Warner, Marvin G; Addleman, R Shane

    2011-08-01

    In the event of an accidental or intentional release of radionuclides into a populated area, massive numbers of people may require radiobioassay screening as triage for dose-reduction therapy or identification for longer-term follow-up. If the event released significant levels of beta- or alpha-emitting radionuclides, in vivo assays would be ineffective. Therefore, highly efficient and rapid analytical methods for radionuclide detection from submitted spot urine samples (≤50 mL) would be required. At present, the quantitative determination of alpha-emitting radionuclides from urine samples is highly labor intensive and requires significant time to prepare and analyze samples. Sorbent materials that provide effective collection and enable rapid assay could significantly streamline the radioanalytical process. The authors have demonstrated the use of magnetic nanoparticles as a novel method of extracting media for four alpha-emitting radionuclides of concern (polonium, radium, uranium and americium) from chemically-unmodified and pH-2 human urine. Herein, the initial experimental sorption results are presented along with a novel method that uses magnetic nanoparticles to extract radionuclides from unmodified human urine and then collect the magnetic field-induced particles for subsequent alpha-counting-source preparation. Additionally, a versatile human dose model is constructed that determines the detector count times required to estimate dose at specific protective-action thresholds. The model provides a means to assess a method's detection capabilities and uses fundamental health physics parameters and actual experimental data as core variables. The modeling shows that, with effective sorbent materials, rapid screening for alpha-emitters is possible with a 50-mL urine sample collected within 1 wk of exposure/intake. PMID:21709509

  16. Magnetic core/shell nanoparticle thin films deposited by MAPLE: Investigation by chemical, morphological and in vitro biological assays

    NASA Astrophysics Data System (ADS)

    Cristescu, R.; Popescu, C.; Socol, G.; Iordache, I.; Mihailescu, I. N.; Mihaiescu, D. E.; Grumezescu, A. M.; Balan, A.; Stamatin, I.; Chifiriuc, C.; Bleotu, C.; Saviuc, C.; Popa, M.; Chrisey, D. B.

    2012-09-01

    We report on thin film deposition of nanostructured Fe3O4/oleic acid/ceftriaxone and Fe3O4/oleic acid/cefepime nanoparticles (core/shell/adsorption-shell) were fabricated by matrix assisted pulsed laser evaporation (MAPLE) onto inert substrates. The thin films were characterized by profilometry, Fourier transform infrared spectroscopy, atomic force microscopy, and investigated by in vitro biological assays. The biological properties tested included the investigation of the microbial viability and the microbial adherence to the glass coverslip nanoparticle film, using Gram-negative and Gram-positive bacterial strains with known antibiotic susceptibility behavior, the microbial adherence to the HeLa cells monolayer grown on the nanoparticle pellicle, and the cytotoxicity on eukaryotic cells. The proposed system, based on MAPLE, could be used for the development of novel anti-microbial materials or strategies for fighting pathogenic biofilms frequently implicated in the etiology of biofilm associated chronic infections.

  17. Competitive fluorescence assay for specific recognition of atrazine by magnetic molecularly imprinted polymer based on Fe3O4-chitosan.

    PubMed

    Liu, Guangyang; Li, Tengfei; Yang, Xin; She, Yongxin; Wang, Miao; Wang, Jing; Zhang, Min; Wang, Shanshan; Jin, Fen; Jin, Maojun; Shao, Hua; Jiang, Zejun; Yu, Hailong

    2016-02-10

    A novel fluorescence sensing strategy for determination of atrazine in tap water involving direct competition between atrazine and 5-(4,6-dichlorotriazinyl) aminofluorescein (5-DTAF), and which exploits magnetic molecularly imprinted polymer (MMIP), has been developed. The MMIP, based on Fe3O4-chitosan nanoparticles, was synthesized to recognize specific binding sites of atrazine. The recognition capability and selectivity of the MMIP for atrazine and other triazine herbicides was investigated. Under optimal conditions, the competitive reaction between 5-DTAF and atrazine was performed to permit quantitation. Fluorescence intensity changes at 515 nm was linearly related to the logarithm of the atrazine concentration for the range 2.32-185.4 μM. The detection limit for atrazine was 0.86μM (S/N=3) and recoveries were 77.6-115% in spiked tap water samples.

  18. Magnetic beads-based enzymatic spectrofluorometric assay for rapid and sensitive detection of antibody against ApxIVA of Actinobacillus pleuropneumoniae.

    PubMed

    Wei, Bo; Li, Fang; Yang, Huicui; Yu, Lei; Zhao, Kaihong; Zhou, Rui; Hu, Yonggang

    2012-05-15

    In this paper, a simple, easily-operated and enzyme-amplified fluorescence immunoassay method using magnetic particles for the detection of antibody against Actinobacillus pleuropneumoniae (APP) has been presented. The A protein of APP Repeats-in-Toxin IV (ApxIVA) with high specificity to the APP species was immobilized onto the magnetic bead surfaces. Horseradish peroxidase (HRP), which can catalyze the substrate 4-hydroxyphenylacetic acid (p-HPA), generating fluorescent bi-p, p'-hydroxyphenylacetic acid (DBDA), was selected as an enzymatic-amplified tracer. The ApxIVA antibody was detected for the presence of APP infection by measuring the fluorescence intensity of DBDA. Under optimal conditions, the calibration plot obtained for standard positive serum was approximately linear within the dilution range 1:160-1:5120. The limit of detection (LOD) for the assay was 1:10240, considerably lower than that of ApxIVA-ELISA (1:320) (S/N=3). A series of repeatability measurements of using 1:320-fold diluted standard positive serum gave reproducible results with a relative standard deviation (RSD) of 4.8% (n=11). The ability of the immunosensor to analyze clinical samples was tested on porcine sera. The immunosensor yielded an efficiency of 89.7%, sensitivity of 90.9% and specificity of 89.3% compared with ApxIVA-ELISA.

  19. Novel circulating peptide biomarkers for esophageal squamous cell carcinoma revealed by a magnetic bead-based MALDI-TOFMS assay

    PubMed Central

    Jia, Kun; Li, Wei; Wang, Feng; Qu, Haixia; Qiao, Yuanyuan; Zhou, Lanping; Sun, Yulin; Ma, Qingwei; Zhao, Xiaohang

    2016-01-01

    Esophageal squamous cell carcinoma (ESCC) is one of the most common malignant neoplasms worldwide. Patients are often diagnosed at advanced stages with poor prognosis due to the absence of obvious early symptoms. Here, we applied a high-throughput serum peptidome analysis to identify circulating peptide markers of ESCC. Weak cationic exchange magnetic beads coupled to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used for two-stage proteotypic peptide profiling in complex serum samples collected from 477 cancer patients and healthy controls. We established a genetic algorithm model containing three significantly differentially expressed peptides at 1,925.5, 2,950.6 and 5,900.0 Da with a sensitivity and specificity of 97.00% and 95.92% in the training set and 97.03% and 100.00% in the validation set, respectively. The model's diagnostic capability was significantly better than SCC-Ag and Cyfra 21–1, especially for early stage ESCC, with an achieved sensitivity of 96.94%. Subsequently, these peptides were identified as fragments of AHSG, TSP1 and FGA by linear ion trap-orbitrap hybrid tandem mass spectrometry. Notably, increased tissue and serum levels of TSP1 in ESCC were verified and correlated with disease progression. In addition, tissue TSP1 was an independent poor prognostic factor in ESCC. In conclusion, the newly established circulating peptide panel and identified proteins could serve as potential biomarkers for the early detection and diagnosis of ESCC. Nevertheless, a larger cohort will be required for further unequivocal validation of their clinical application. PMID:26993605

  20. Novel circulating peptide biomarkers for esophageal squamous cell carcinoma revealed by a magnetic bead-based MALDI-TOFMS assay.

    PubMed

    Jia, Kun; Li, Wei; Wang, Feng; Qu, Haixia; Qiao, Yuanyuan; Zhou, Lanping; Sun, Yulin; Ma, Qingwei; Zhao, Xiaohang

    2016-04-26

    Esophageal squamous cell carcinoma (ESCC) is one of the most common malignant neoplasms worldwide. Patients are often diagnosed at advanced stages with poor prognosis due to the absence of obvious early symptoms. Here, we applied a high-throughput serum peptidome analysis to identify circulating peptide markers of ESCC. Weak cationic exchange magnetic beads coupled to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used for two-stage proteotypic peptide profiling in complex serum samples collected from 477 cancer patients and healthy controls. We established a genetic algorithm model containing three significantly differentially expressed peptides at 1,925.5, 2,950.6 and 5,900.0 Da with a sensitivity and specificity of 97.00% and 95.92% in the training set and 97.03% and 100.00% in the validation set, respectively. The model's diagnostic capability was significantly better than SCC-Ag and Cyfra 21-1, especially for early stage ESCC, with an achieved sensitivity of 96.94%. Subsequently, these peptides were identified as fragments of AHSG, TSP1 and FGA by linear ion trap-orbitrap hybrid tandem mass spectrometry. Notably, increased tissue and serum levels of TSP1 in ESCC were verified and correlated with disease progression. In addition, tissue TSP1 was an independent poor prognostic factor in ESCC. In conclusion, the newly established circulating peptide panel and identified proteins could serve as potential biomarkers for the early detection and diagnosis of ESCC. Nevertheless, a larger cohort will be required for further unequivocal validation of their clinical application. PMID:26993605

  1. In-syringe magnetic stirring-assisted dispersive liquid-liquid microextraction for automation and downscaling of methylene blue active substances assay.

    PubMed

    Suárez, Ruth; Horstkotte, Burkhard; Cerdà, Victor

    2014-12-01

    A simple and rapid method for the determination of the methylene blue active substances assay based on in-syringe automation of magnetic stirring-assisted dispersive liquid-liquid microextraction was developed. The proposed method proved to be valid for the determination of anionic surfactant in waste, pond, well, tap, and drinking water samples. Sample mixing with reagents, extraction and phase separation were performed within the syringe of an automated syringe pump containing a magnetic stirring bar for homogenization and solvent dispersion. The syringe module was used upside-down to enable the use of chloroform as an extraction solvent of higher density than water. The calibration was found to be linear up to 0.3mg/L using only 200 µL of solvent and 4 mL of sample. The limits of detection (3σ) and quantification (10σ) were 7.0 µg/L and 22 µg/L, respectively. The relative standard deviation for 10 replicate determinations of 0.1mg/L SBDS was below 3%. Concentrations of anionic surfactants in natural water samples were in the range of 0.032-0.213 mg/L and no significant differences towards the standard method were found. Standard additions gave analyte recoveries between 95% and 106% proving the general applicability and adequateness of the system to MBSA index determination. Compared to the tedious standard method requiring up to 50 mL of chloroform, the entire procedure took only 345 s using 250-times less solvent.

  2. A simple method for the identification and assay of iopamidol and iothalamate meglumine in pharmaceutical samples based on proton nuclear magnetic resonance spectroscopy.

    PubMed

    Hanna, G M; Lau-Cam, C A

    2001-02-01

    A proton nuclear magnetic resonance (PMR) spectroscopic method is described for the direct assay and identification of the triiodinated radiographic contrast agents iopamidol (nonionic type) and iothalamate meglumine (ionic type) in commercial solutions and as a bulk material. Samples were prepared by simply diluting an injectable solution with or dissolving a powdered sample in D2O. Sodium acetate was added to serve as an internal standard. Quantitations were based on the resonance signals for the protons of the CH3-CO-group at 1.58 ppm (iopamidol) or 2.25 ppm (iothalamate), CH3-N-group at 2.38 ppm (meglumine) and CH3-CO-group at 1.92 ppm (acetate). The mean +/- SD (n = sets of 10 samples each) recovery of iopamidol, iothalamic acid and meglumine from synthetic mixtures with the internal standard were 99.6 +/- 0.63, 99.7 +/- 0.66 and 99.9 +/- 1.18%, respectively; with the values ranging from 98.7-100.9% for iopamidol and iothalamate, and from 98.3-100.8% for meglumine.

  3. Investigation of magnetic nanoparticles for the rapid extraction and assay of alpha-emitting radionuclides from urine: Demonstration of a novel radiobioassay method

    SciTech Connect

    O'Hara, Matthew J.; Carter, Jennifer C.; Maclellan, Jay A.; Warner, Cynthia L.; Warner, Marvin G.; Addleman, Raymond S.

    2011-08-01

    In the event of an accidental or intentional release of radionuclides into a populated area, three things must occur in a timely manner: food and drinking water supplies must be determined to be safe to eat / drink, civilians and/or military personnel must be surveyed to ensure that they do not have external contamination, and they must be screened to ensure that significant ingestion or inhalation of radionuclides has not occurred (this paper is concerned with the latter). In the event of such a disaster, the volume of radiobioassays to be performed would be tremendous. If the event released significant levels of β- or α-emitting radionuclides, in vivo assays would be ineffective. Therefore, highly efficient and rapid analytical methods for radionuclide detection from submitted spot urine samples (≤ 50 mL) would be required. At present, the quantitative determination of α-emitting radionuclides from urine samples is highly labor intensive, and requires significant sample preparation and analysis time. Sorbent materials that provide effective collection and enable rapid assay could significantly streamline the radioanalytical process. We have demonstrated the use of paramagnetic nanoparticles as a novel class of extracting media for four α-emitting radionuclides of concern (Po, Ra, Am, and U) from chemically unmodified and pH 2 human urine. Herein the initial experimental sorption results are presented along with a novel method that utilizes paramagnetic nanoparticles for the extraction of radionuclides from unmodified human urine followed by the magnetic field-induced collection of the particles for subsequent α-counting-source preparation. Additionally, we construct a versatile human dose model that determines the detector count times required to estimate internal human dose at specific protective action thresholds. The model provides a means to assess a method’s detection capabilities and use fundamental health physics parameters and actual experimental

  4. Evaluating the influence of National Research Council levels of copper, iron, manganese, and zinc using organic (Bioplex) minerals on resulting tissue mineral concentrations, metallothionein, and liver antioxidant enzymes in grower-finisher swine diets.

    PubMed

    Gowanlock, D W; Mahan, D C; Jolliff, J S; Hill, G M

    2015-03-01

    Graded levels of a trace mineral premix containing an organic (Bioplex) source of Cu, Fe, Mn, and Zn was evaluated with additional treatments containing organic Zn or Fe. Grower-finisher pigs were fed from 25 to 115 kg BW. The number of pigs in the experiment, the breeding/genetics of the pigs, the management, and the average age of the pigs were previously reported. The experiment was conducted as a randomized complete block design in 7 replicates. Treatments were 1) basal diet without supplemental Cu, Fe, Mn, and Zn; 2) basal diet + 2.5 mg/kg Cu, 50 mg/kg Fe, 1.5 mg/kg Mn, and 40 mg/kg Zn (50% NRC); 3) basal diet + 5 mg/kg Cu, 100 mg/kg Fe, 3 mg/kg Mn, and 80 mg/kg Zn (100% NRC); 4) basal diet + 25 mg Zn/kg; 5) basal diet + 50 mg Zn/kg; and 6) basal diet + 50 mg Fe/kg. Selenium and I were added to all diets at 0.3 and 0.14 mg/kg, respectively. Diets were composed of corn-soybean meal, dicalcium phosphate, and limestone with phytase added to enhance mineral availability. Three pigs per pen were bled at 55, 80, and 115 kg BW and plasma was analyzed for microminerals. When the average replicate BW was 115 kg, 3 pigs per pen of an equal gender ratio were killed. The liver, kidney, and heart were removed and analyzed for microminerals. Liver, duodenum, and jejunal metallothionein and the antioxidant enzymes in the liver containing these microminerals were determined. The results demonstrated that plasma minerals were unaffected at the 3 BW intervals. Liver and duodenum metallothionein protein were greater ( < 0.05) as dietary micromineral levels increased but jejunum metallothionein did not change as microminerals increased. The activity of Cu/Zn superoxide dismutase (SOD) was not affected as the levels of the micromineral increased, whereas the activity of Mn SOD increased slightly ( < 0.05) to the 50% NRC treatment level. Liver Zn (relative and total) increased ( < 0.05) as dietary micromineral levels increased and also when Zn was added singly to the diet. Liver

  5. Cellulase Assays

    NASA Astrophysics Data System (ADS)

    Zhang, Y. H. Percival; Hong, Jiong; Ye, Xinhao

    Cellulose is a heterogeneous polysaccharide, and its enzymatic hydrolysis requires endoglucanase, exoglucanase (cellobiohydrolase), and β-glucosidase to work together. We summarize the most commonly used assays for individual enzymes and cellulase mixture.

  6. Treponema-specific tests for serodiagnosis of syphilis: comparative evaluation of seven assays.

    PubMed

    Binnicker, M J; Jespersen, D J; Rollins, L O

    2011-04-01

    The diagnosis of syphilis is challenging and often relies on serologic tests to detect treponemal or nontreponemal antibodies. Recently, the Centers for Disease Control and Prevention and the Association of Public Health Laboratories proposed an update to the syphilis serology testing algorithm, in which serum samples are first tested using a treponema-specific test and positive samples are analyzed with a nontreponemal assay. The goal of this study was to compare the performance of seven treponemal assays (BioPlex 2200 syphilis IgG [Bio-Rad, Hercules, CA], fluorescent treponemal antibody [FTA] assay [Zeus Scientific, Raritan, NJ], Treponema pallidum particle agglutination [TP-PA; Fujirebio Diagnostics, Malvern, PA], Trep-Sure enzyme immunoassay [EIA; Phoenix Biotech, Oakville, Ontario, Canada], Trep-Chek EIA [Phoenix Biotech], Trep-ID EIA [Phoenix Biotech], and Treponema ViraBlot IgG [Viramed Biotech AG, Planegg, Germany]) using serum samples (n = 303) submitted to our reference laboratory. In addition to testing with these 7 assays, all samples were tested by a rapid plasma reagin (RPR) assay and a treponemal IgM Western blot assay (Viramed ViraBlot). Compared to the FTA assay as the gold standard, the evaluated treponemal tests demonstrated comparable levels of performance, with percent agreement ranging from 95.4% (95% confidence interval, 92.3 to 97.3) for the Trep-Sure EIA to 98.4% (96.1 to 99.4) for the Trep-ID EIA. Compared to a "consensus of the test panel" (defined as at least 4 of 7 treponemal tests being in agreement), the percent agreement ranged from 95.7% (92.7 to 97.5) for Trep-Sure to 99.3% (97.5 to 99.9) for Trep-ID. These data may assist clinical laboratories that are considering implementing a treponemal test for screening or confirmatory purposes.

  7. MAGNETS

    DOEpatents

    Hofacker, H.B.

    1958-09-23

    This patent relates to nmgnets used in a calutron and more particularly to means fur clamping an assembly of magnet coils and coil spacers into tightly assembled relation in a fluid-tight vessel. The magnet comprises windings made up of an assembly of alternate pan-cake type coils and spacers disposed in a fluid-tight vessel. At one end of the tank a plurality of clamping strips are held firmly against the assembly by adjustable bolts extending through the adjacent wall. The foregoing arrangement permits taking up any looseness which may develop in the assembly of coils and spacers.

  8. Highly specific and rapid immuno-fluorescent visualization and detection of E. coli O104:H4 with protein-A coated magnetic beads based LST-MUG assay.

    PubMed

    Barizuddin, Syed; Balakrishnan, Baskar; Stringer, R Cody; Dweik, Majed

    2015-08-01

    A method combining immunomagnetic separation and fluorescent sensing was developed to detect Escherichia coli (E. coli) O104:H4. The antibody specific to E. coli O104:H4 was immobilized on protein A-coated magnetic beads. This protein-A-anti E. coli O104:H4 complex was used to bind Fluorescein IsoThioCyanate (FITC) labeled E. coli O104:H4 antigen (whole cell) on it. The goal was to achieve a fluorescently detectable protein-A-anti E. coli O104:H4-E. coli O104:H4 complex on the magnetic beads. Fluorescent microscopy was used to image the magnetic beads. The resulting fluorescence on the beads was due to the FITC labeled antigen binding on the protein-A-anti E. coli O104:H4 immobilized magnetic beads. This visually proves the antigen-antibody binding. The fluorescent imaging results were obtained in 2 h if the minimum available bacteria in the sample were at least 10(5) CFU/ml. If no fluorescence was observed on the magnetic beads during fluorescent imaging, it indicates the bacterial concentration in the sample to be too low for it to have bound to the magnetic beads and hence no detection was possible. To detect bacterial concentration less than 10(5) CFU/ml in the sample, an additional step was required for detection. The magnetic bead complex was added to the LST-MUG (lauryl sulfate tryptose-4-methylumbelliferyl-β-D-glucuronide), a signaling reporter. The E. coli O104:H4 grows in LST-MUG and releases β-glucuronidase enzyme. This enzyme cleaves the MUG substrate that produces 4-methylumbelliferone, a highly fluorescent species. This fluorescence was detected using a spectrofluorometer. The emission peak in the fluorescent spectrum was found to be at 450 nm. The lower and upper detection range for this LST-MUG assay was found to be 2.05×10(5)-4.09×10(8) CFU/ml. The results for the LST-MUG assay for concentrations below 10(5) CFU/ml were ascertained in 8h. The advantages of this technique include the specific detection of bacteria without an enrichment step and

  9. Environmental Technology Verification Report for Abraxis 17β-Estradiol (E2) Magnetic Particle Enzyme-Linked Immunosorbent Assay (ELISA) Test Kits

    EPA Science Inventory

    The EPA's National Risk Management Research Laboratory (NRMRL) and its verification organization partner, Battelle, operate the Advanced Monitoring Systems (AMS) Center under ETV. The AMS Center recently evaluated the performance of the Abraxis 17(beta)-estradiol (E2) magnetic p...

  10. Independency of Fe ions in hemoglobin on immunomagnetic reduction assay

    NASA Astrophysics Data System (ADS)

    Yang, S. Y.; Lan, C. B.; Chen, C. H.; Horng, H. E.; Hong, Chin-Yih; Yang, H. C.; Lai, Y. K.; Lin, Y. H.; Teng, K. S.

    2009-10-01

    Immunomagnetic reduction (IMR), which involves measuring the reduction in the ac magnetic susceptibility of magnetic reagents, is due to the association between bio-functionalized magnetic nanoparticles and target bio-molecules. This has been demonstrated for assaying proteins in solutions free of Fe ions, such as serum. In this work, the validity of IMR assay for samples rich in Fe ions like hemoglobin (Hb) is investigated. According to the results, there is no magnetic signal contributed by Fe-ion-rich Hb. Furthermore, the results show a high sensitivity in assaying hemoglobin A1c (HbA1c) by using IMR.

  11. Magnetic Nanoparticle Sensors

    PubMed Central

    Koh, Isaac; Josephson, Lee

    2009-01-01

    Many types of biosensors employ magnetic nanoparticles (diameter = 5–300 nm) or magnetic particles (diameter = 300–5,000 nm) which have been surface functionalized to recognize specific molecular targets. Here we cover three types of biosensors that employ different biosensing principles, magnetic materials, and instrumentation. The first type consists of magnetic relaxation switch assay-sensors, which are based on the effects magnetic particles exert on water proton relaxation rates. The second type consists of magnetic particle relaxation sensors, which determine the relaxation of the magnetic moment within the magnetic particle. The third type is magnetoresistive sensors, which detect the presence of magnetic particles on the surface of electronic devices that are sensitive to changes in magnetic fields on their surface. Recent improvements in the design of magnetic nanoparticles (and magnetic particles), together with improvements in instrumentation, suggest that magnetic material-based biosensors may become widely used in the future. PMID:22408498

  12. Magnetic nanotubes

    DOEpatents

    Matsui, Hiroshi; Matsunaga, Tadashi

    2010-11-16

    A magnetic nanotube includes bacterial magnetic nanocrystals contacted onto a nanotube which absorbs the nanocrystals. The nanocrystals are contacted on at least one surface of the nanotube. A method of fabricating a magnetic nanotube includes synthesizing the bacterial magnetic nanocrystals, which have an outer layer of proteins. A nanotube provided is capable of absorbing the nanocrystals and contacting the nanotube with the nanocrystals. The nanotube is preferably a peptide bolaamphiphile. A nanotube solution and a nanocrystal solution including a buffer and a concentration of nanocrystals are mixed. The concentration of nanocrystals is optimized, resulting in a nanocrystal to nanotube ratio for which bacterial magnetic nanocrystals are immobilized on at least one surface of the nanotubes. The ratio controls whether the nanocrystals bind only to the interior or to the exterior surfaces of the nanotubes. Uses include cell manipulation and separation, biological assay, enzyme recovery, and biosensors.

  13. RAS - Screens & Assays

    Cancer.gov

    A primary goal of the RAS Initiative is to develop assays for RAS activity, localization, and signaling and adapt those assays so they can be used for finding new drug candidates. Explore the work leading to highly validated screening protocols.

  14. Mass Spectrometry and Multiplex Antigen Assays to Assess Microbial Quality and Toxin Production of Staphylococcus aureus Strains Isolated from Clinical and Food Samples

    PubMed Central

    Attien, Paul; Sina, Haziz; Moussaoui, Wardi; Zimmermann-Meisse, Gaëlle; Dadié, Thomas; Keller, Daniel; Riegel, Philippe; Edoh, Vincent; Kotchoni, Simeon O.; Djè, Marcellin; Prévost, Gilles

    2014-01-01

    The aim of our study was to investigate the microbial quality of meat products and on some clinical samples in Abidjan focused on Staphylococcus genus and the toxin production profile of Staphylococcus aureus (S. aureus) isolated. Bacteria were collected from 240 samples of three meat products sold in Abidjan and 180 samples issued from clinical infections. The strains were identified by both microbiological and MALDI-TOF-MS methods. The susceptibility to antibiotics was determined by the disc diffusion method. The production of Panton-Valentine Leukocidin, LukE/D, and epidermolysins was screened using radial gel immunodiffusion. The production of staphylococcal enterotoxins and TSST-1 was screened by a Bio-Plex Assay. We observed that 96/240 of meat samples and 32/180 of clinical samples were contaminated by Staphylococcus. Eleven species were isolated from meats and 4 from clinical samples. Forty-two S. aureus strains were isolated from ours samples. Variability of resistance was observed for most of the tested antibiotics but none of the strains displays a resistance to imipenem and quinolones. We observed that 89% of clinical S. aureus were resistant to methicillin against 58% for those issued from meat products. All S. aureus isolates issued from meat products produce epidermolysins whereas none of the clinical strains produced these toxins. The enterotoxins were variably produced by both clinical and meat product samples. PMID:24987686

  15. Rotor assembly and assay method

    DOEpatents

    Burtis, Carl A.; Johnson, Wayne F.; Walker, William A.

    1993-01-01

    A rotor assembly for carrying out an assay includes a rotor body which is rotatable about an axis of rotation, and has a central chamber and first, second, third, fourth, fifth, and sixth chambers which are in communication with and radiate from the central chamber. The rotor assembly further includes a shuttle which is movable through the central chamber and insertable into any of the chambers, the shuttle including a reaction cup carrying an immobilized antigen or an antibody for transport among the chambers. A method for carrying out an assay using the rotor assembly includes moving the reaction cup among the six chambers by passing the cup through the central chamber between centrifugation steps in order to perform the steps of: separating plasma from blood cells, binding plasma antibody or antigen, washing, drying, binding enzyme conjugate, reacting with enzyme substrate and optically comparing the resulting reaction product with unreacted enzyme substrate solution. The movement of the reaction cup can be provided by attaching a magnet to the reaction cup and supplying a moving magnetic field to the rotor.

  16. Rotor assembly and assay method

    DOEpatents

    Burtis, C.A.; Johnson, W.F.; Walker, W.A.

    1993-09-07

    A rotor assembly for carrying out an assay includes a rotor body which is rotatable about an axis of rotation, and has a central chamber and first, second, third, fourth, fifth, and sixth chambers which are in communication with and radiate from the central chamber. The rotor assembly further includes a shuttle which is movable through the central chamber and insertable into any of the chambers, the shuttle including a reaction cup carrying an immobilized antigen or an antibody for transport among the chambers. A method for carrying out an assay using the rotor assembly includes moving the reaction cup among the six chambers by passing the cup through the central chamber between centrifugation steps in order to perform the steps of: separating plasma from blood cells, binding plasma antibody or antigen, washing, drying, binding enzyme conjugate, reacting with enzyme substrate and optically comparing the resulting reaction product with unreacted enzyme substrate solution. The movement of the reaction cup can be provided by attaching a magnet to the reaction cup and supplying a moving magnetic field to the rotor. 34 figures.

  17. Absolute nuclear material assay

    DOEpatents

    Prasad, Manoj K.; Snyderman, Neal J.; Rowland, Mark S.

    2012-05-15

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  18. Absolute nuclear material assay

    DOEpatents

    Prasad, Manoj K.; Snyderman, Neal J.; Rowland, Mark S.

    2010-07-13

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  19. CPTAC Assay Portal: a repository of targeted proteomic assays

    SciTech Connect

    Whiteaker, Jeffrey R.; Halusa, Goran; Hoofnagle, Andrew N.; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John; Kennedy, Jacob; Mani, DR; Zimmerman, Lisa J.; Meyer, Matthew R.; Mesri, Mehdi; Rodriguez, Henry; Abbateillo, Susan E.; Boja, Emily; Carr, Steven A.; Chan, Daniel W.; Chen, Xian; Chen, Jing; Davies, Sherri; Ellis, Matthew; Fenyo, David; Hiltket, Tara; Ketchum, Karen; Kinsinger, Christopher; Kuhn, Eric; Liebler, Daniel; Lin, De; Liu, Tao; Loss, Michael; MacCoss, Michael; Qian, Weijun; Rivers, Robert; Rodland, Karin D.; Ruggles, Kelly; Scott, Mitchell; Smith, Richard D.; Thomas, Stefani N.; Townsend, Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Paulovich, Amanda G.

    2014-06-27

    To address these issues, the Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal (http://assays.cancer.gov) to serve as a public repository of well-characterized quantitative, MS-based, targeted proteomic assays. The purpose of the CPTAC Assay Portal is to facilitate widespread adoption of targeted MS assays by disseminating SOPs, reagents, and assay characterization data for highly characterized assays. A primary aim of the NCI-supported portal is to bring together clinicians or biologists and analytical chemists to answer hypothesis-driven questions using targeted, MS-based assays. Assay content is easily accessed through queries and filters, enabling investigators to find assays to proteins relevant to their areas of interest. Detailed characterization data are available for each assay, enabling researchers to evaluate assay performance prior to launching the assay in their own laboratory.

  20. Lateral flow assays.

    PubMed

    Koczula, Katarzyna M; Gallotta, Andrea

    2016-06-30

    Lateral flow assays (LFAs) are the technology behind low-cost, simple, rapid and portable detection devices popular in biomedicine, agriculture, food and environmental sciences. This review presents an overview of the principle of the method and the critical components of the assay, focusing on lateral flow immunoassays. This type of assay has recently attracted considerable interest because of its potential to provide instantaneous diagnosis directly to patients. The range and interpretation of results and parameters used for evaluation of the assay will also be discussed. The main advantages and disadvantages of LFAs will be summarized and relevant future improvements to testing devices and strategies will be proposed. Finally, the major recent advances and future diagnostic applications in the LFA field will be explored. PMID:27365041

  1. Lateral flow assays

    PubMed Central

    Koczula, Katarzyna M.

    2016-01-01

    Lateral flow assays (LFAs) are the technology behind low-cost, simple, rapid and portable detection devices popular in biomedicine, agriculture, food and environmental sciences. This review presents an overview of the principle of the method and the critical components of the assay, focusing on lateral flow immunoassays. This type of assay has recently attracted considerable interest because of its potential to provide instantaneous diagnosis directly to patients. The range and interpretation of results and parameters used for evaluation of the assay will also be discussed. The main advantages and disadvantages of LFAs will be summarized and relevant future improvements to testing devices and strategies will be proposed. Finally, the major recent advances and future diagnostic applications in the LFA field will be explored. PMID:27365041

  2. Tube-Forming Assays.

    PubMed

    Brown, Ryan M; Meah, Christopher J; Heath, Victoria L; Styles, Iain B; Bicknell, Roy

    2016-01-01

    Angiogenesis involves the generation of new blood vessels from the existing vasculature and is dependent on many growth factors and signaling events. In vivo angiogenesis is dynamic and complex, meaning assays are commonly utilized to explore specific targets for research into this area. Tube-forming assays offer an excellent overview of the molecular processes in angiogenesis. The Matrigel tube forming assay is a simple-to-implement but powerful tool for identifying biomolecules involved in angiogenesis. A detailed experimental protocol on the implementation of the assay is described in conjunction with an in-depth review of methods that can be applied to the analysis of the tube formation. In addition, an ImageJ plug-in is presented which allows automatic quantification of tube images reducing analysis times while removing user bias and subjectivity.

  3. New Rapid Spore Assay

    NASA Astrophysics Data System (ADS)

    Kminek, Gerhard; Conley, Catharine

    2012-07-01

    The presentation will detail approved Planetary Protection specifications for the Rapid Spore Assay for spacecraft components and subsystems. Outlined will be the research and studies on which the specifications were based. The research, funded by ESA and NASA/JPL, was conducted over a period of two years and was followed by limited cleanroom studies to assess the feasibility of this assay during spacecraft assembly.

  4. Doped colorimetric assay liposomes

    DOEpatents

    Charych, Deborah; Stevens, Raymond C.

    2001-01-01

    The present invention provides compositions comprising colorimetric assay liposomes. The present invention also provides methods for producing colorimetric liposomes and calorimetric liposome assay systems. In preferred embodiments, these calorimetric liposome systems provide high levels of sensitivity through the use of dopant molecules. As these dopants allow the controlled destabilization of the liposome structure, upon exposure of the doped liposomes to analyte(s) of interest, the indicator color change is facilitated and more easily recognized.

  5. SNAP Assay Technology.

    PubMed

    O'Connor, Thomas P

    2015-12-01

    The most widely used immunoassay configuration is the enzyme-linked immunosorbent assay (ELISA) because the procedure produces highly sensitive and specific results and generally is easy to use. By definition, ELISAs are immunoassays used to detect a substance (typically an antigen or antibody) in which an enzyme is attached (conjugated) to one of the reactants and an enzymatic reaction is used to amplify the signal if the substance is present. Optimized ELISAs include several steps that are performed in sequence using a defined protocol that typically includes application of sample and an enzyme-conjugated antibody or antigen to an immobilized reagent, followed by wash and enzyme reaction steps. The SNAP assay is an in-clinic device that performs each of the ELISA steps in a timed sequential fashion with little consumer interface. The components and mechanical mechanism of the assay device are described. Detailed descriptions of features of the assay, which minimize nonspecific binding and enhance the ability to read results from weak-positive samples, are given. Basic principles used in assays with fundamentally different reaction mechanisms, namely, antigen-detection, antibody-detection, and competitive assays are given. Applications of ELISA technology, which led to the development of several multianalyte SNAP tests capable of testing for up to 6 analytes using a single-sample and a single-SNAP device are described.

  6. Against vaccine assay secrecy.

    PubMed

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M

    2015-01-01

    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors.

  7. Rover waste assay system

    SciTech Connect

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J.

    1997-11-01

    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched {sup 235}U contaminated piping, tank sections, and debris from the Rover nuclear rocket fuel processing facility at the Idaho Chemical Processing Plant. A scanning system translates a NaI(Tl) detector/collimator system over the structural components where both relative and calibrated measurements for {sup 137}Cs are made. Uranium-235 concentrations are in operation and is sufficiently automated that most functions are performed by the computer system. These functions include system calibration, problem identification, collimator control, data analysis, and reporting. Calibration of the system was done through a combination of measurements on calibration standards and benchmarked modeling. A description of the system is presented along with the methods and uncertainties associated with the calibration and analysis of the system for components from the Rover facility. 4 refs., 2 figs., 4 tabs.

  8. Interpreting coagulation assays.

    PubMed

    Green, David

    2010-09-01

    The interpretation of coagulation assays requires knowledge of the principal clotting pathways. The activated partial thromboplastin time is sensitive to all hemostatic factors except FVII, whereas the prothrombin time reflects levels of prothrombin and FV, FVII, and FX. Using the two tests in concert is helpful in identifying hemophilia, the coagulopathy of liver disease, and disseminated intravascular coagulation. In addition, the activated partial thromboplastin time and prothrombin time are used for monitoring anticoagulant therapy with heparin and warfarin, respectively. Measurement of D-dimer is informative in patients suspected of having thrombotic disorders and determining the risk of thrombosis recurrence. Mixing tests distinguish clotting factor deficiencies from circulating anticoagulants such as heparin, the lupus anticoagulant, and antibodies directed against specific clotting factors. The modified Bethesda assay detects and provides an indication of the strength of FVIII inhibitors. However, interpreting the results of coagulation assays is not always straightforward, and expert consultation is occasionally required to resolve difficult clinical situations. PMID:20855988

  9. Against vaccine assay secrecy

    PubMed Central

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M

    2015-01-01

    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors. PMID:25826194

  10. Multiplex Flow Assays

    PubMed Central

    2016-01-01

    Lateral flow or dipstick assays (e.g., home pregnancy tests), where an analyte solution is drawn through a porous membrane and is detected by localization onto a capture probe residing at a specific site on the flow strip, are the most commonly and extensively used type of diagnostic assay. However, after over 30 years of use, these assays are constrained to measuring one or a few analytes at a time. Here, we describe a completely general method, in which any single-plex lateral flow assay is transformed into a multiplex assay capable of measuring an arbitrarily large number of analytes simultaneously. Instead of identifying the analyte by its localization onto a specific geometric location in the flow medium, the analyte-specific capture probe is identified by its association with a specific optically encoded region within the flow medium. The capture probes for nucleic acids, antigens, or antibodies are attached to highly porous agarose beads, which have been encoded using multiple lanthanide emitters to create a unique optical signature for each capture probe. The optically encoded capture probe-derivatized beads are placed in contact with the analyte-containing porous flow medium and the analytes are captured onto the encoded regions as the solution flows through the porous medium. To perform a multiplex diagnostic assay, a solution comprising multiple analytes is passed through the flow medium containing the capture probe-derivatized beads, and the captured analyte is treated with a suitable fluorescent reporter. We demonstrate this multiplex analysis technique by simultaneously measuring DNA samples, antigen–antibody pairs, and mixtures of multiple nucleic acids and antibodies.

  11. Fluorometric assay for aflatoxins

    SciTech Connect

    Chakrabarti, A.G.

    1984-11-01

    The method that is now widely adopted by the government laboratories for the assay of individual aflatoxin components (B/sub 1/, B/sub 2/, G/sub 1/, and G/sub 2/) utilizes a TLC technique. The extraction and clean-up steps of this technique were further researched but the method is still time consuming. It is, therefore, very important to develop a rapid and accurate assay technique for aflatoxins. The current research proposes a technique which utilizes a Turner Fluorometer.

  12. Lateral flow strip assay

    DOEpatents

    Miles, Robin R.; Benett, William J.; Coleman, Matthew A.; Pearson, Francesca S.; Nasarabadi, Shanavaz L.

    2011-03-08

    A lateral flow strip assay apparatus comprising a housing; a lateral flow strip in the housing, the lateral flow strip having a receiving portion; a sample collection unit; and a reagent reservoir. Saliva and/or buccal cells are collected from an individual using the sample collection unit. The sample collection unit is immersed in the reagent reservoir. The tip of the lateral flow strip is immersed in the reservoir and the reagent/sample mixture wicks up into the lateral flow strip to perform the assay.

  13. Instrument for assaying radiation

    DOEpatents

    Coleman, Jody Rustyn; Farfan, Eduardo B.

    2016-03-22

    An instrument for assaying radiation includes a flat panel detector having a first side opposed to a second side. A collimated aperture covers at least a portion of the first side of the flat panel detector. At least one of a display screen or a radiation shield may cover at least a portion of the second side of the flat panel detector.

  14. Kinetic tetrazolium microtiter assay

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L. (Inventor); Stowe, Raymond P. (Inventor); Koeing, David W. (Inventor)

    1992-01-01

    A method for conducting an in vitro cell assay using a tetrazolium indicator is disclosed. The indicator includes a nonionic detergent which solubilizes a tetrazolium reduction product in vitro and has low toxicity for the cells. The incubation of test cells in the presence of zolium bromide and octoxynol (TRITON X-100) permits kinetics of the cell metabolism to be determined.

  15. Kinetic Tetrazolium Microtiter Assay

    NASA Technical Reports Server (NTRS)

    Pierson, Duane L.; Stowe, Raymond; Koenig, David

    1993-01-01

    Kinetic tetrazolium microtiter assay (KTMA) involves use of tetrazolium salts and Triton X-100 (or equivalent), nontoxic, in vitro color developer solubilizing colored metabolite formazan without injuring or killing metabolizing cells. Provides for continuous measurement of metabolism and makes possible to determine rate of action of antimicrobial agent in real time as well as determines effective inhibitory concentrations. Used to monitor growth after addition of stimulatory compounds. Provides for kinetic determination of efficacy of biocide, greatly increasing reliability and precision of results. Also used to determine relative effectiveness of antimicrobial agent as function of time. Capability of generating results on day of test extremely important in treatment of water and waste, disinfection of hospital rooms, and in pharmaceutical, agricultural, and food-processing industries. Assay also used in many aspects of cell biology.

  16. Radioreceptor assay for oxyphenonium.

    PubMed

    Ensing, K; de Zeeuw, R A

    1984-01-01

    The development of a radioreceptor assay for the quaternary anticholinergic drug, oxyphenonium, in plasma is reported. It is based on competition between this drug and 3H-dexetimide for binding to muscarinic receptors. After ion pair extraction and reextraction, the drug can be determined in plasma at concentrations down to a value of 100 pg/ml. This permits pharmacokinetic studies to be made after inhalation of oxyphenonium. PMID:6428927

  17. C. elegans chemotaxis assay.

    PubMed

    Margie, Olivia; Palmer, Chris; Chin-Sang, Ian

    2013-01-01

    Many organisms use chemotaxis to seek out food sources, avoid noxious substances, and find mates. Caenorhabditis elegans has impressive chemotaxis behavior. The premise behind testing the response of the worms to an odorant is to place them in an area and observe the movement evoked in response to an odorant. Even with the many available assays, optimizing worm starting location relative to both the control and test areas, while minimizing the interaction of worms with each other, while maintaining a significant sample size remains a work in progress (1-10). The method described here aims to address these issues by modifying the assay developed by Bargmann et al.(1). A Petri dish is divided into four quadrants, two opposite quadrants marked "Test" and two are designated "Control". Anesthetic is placed in all test and control sites. The worms are placed in the center of the plate with a circle marked around the origin to ensure that non-motile worms will be ignored. Utilizing a four-quadrant system rather than one 2 or two 1 eliminates bias in the movement of the worms, as they are equidistant from test and control samples, regardless of which side of the origin they began. This circumvents the problem of worms being forced to travel through a cluster of other worms to respond to an odorant, which can delay worms or force them to take a more circuitous route, yielding an incorrect interpretation of their intended path. This method also shows practical advantages by having a larger sample size and allowing the researcher to run the assay unattended and score the worms once the allotted time has expired. PMID:23644543

  18. Metal-amplified Density Assays, (MADAs), including a Density-Linked Immunosorbent Assay (DeLISA).

    PubMed

    Subramaniam, Anand Bala; Gonidec, Mathieu; Shapiro, Nathan D; Kresse, Kayleigh M; Whitesides, George M

    2015-02-21

    This paper reports the development of Metal-amplified Density Assays, or MADAs - a method of conducting quantitative or multiplexed assays, including immunoassays, by using Magnetic Levitation (MagLev) to measure metal-amplified changes in the density of beads labeled with biomolecules. The binding of target analytes (i.e. proteins, antibodies, antigens) to complementary ligands immobilized on the surface of the beads, followed by a chemical amplification of the binding in a form that results in a change in the density of the beads (achieved by using gold nanoparticle-labeled biomolecules, and electroless deposition of gold or silver), translates analyte binding events into changes in density measureable using MagLev. A minimal model based on diffusion-limited growth of hemispherical nuclei on a surface reproduces the dynamics of the assay. A MADA - when performed with antigens and antibodies - is called a Density-Linked Immunosorbent Assay, or DeLISA. Two immunoassays provided a proof of principle: a competitive quantification of the concentration of neomycin in whole milk, and a multiplexed detection of antibodies against Hepatitis C virus NS3 protein and syphilis T. pallidum p47 protein in serum. MADAs, including DeLISAs, require, besides the requisite biomolecules and amplification reagents, minimal specialized equipment (two permanent magnets, a ruler or a capillary with calibrated length markings) and no electrical power to obtain a quantitative readout of analyte concentration. With further development, the method may be useful in resource-limited or point-of-care settings. PMID:25474561

  19. Radon assay for SNO+

    SciTech Connect

    Rumleskie, Janet

    2015-12-31

    The SNO+ experiment will study neutrinos while located 6,800 feet below the surface of the earth at SNOLAB. Though shielded from surface backgrounds, emanation of radon radioisotopes from the surrounding rock leads to back-grounds. The characteristic decay of radon and its daughters allows for an alpha detection technique to count the amount of Rn-222 atoms collected. Traps can collect Rn-222 from various positions and materials, including an assay skid that will collect Rn-222 from the organic liquid scintillator used to detect interactions within SNO+.

  20. Growth cone collapse assay.

    PubMed

    Cook, Geoffrey M W; Jareonsettasin, Prem; Keynes, Roger J

    2014-01-01

    The growth cone collapse assay has proved invaluable in detecting and purifying axonal repellents. Glycoproteins/proteins present in detergent extracts of biological tissues are incorporated into liposomes, added to growth cones in culture and changes in morphology are then assessed. Alternatively purified or recombinant molecules in aqueous solution may be added directly to the cultures. In both cases after a defined period of time (up to 1 h), the cultures are fixed and then assessed by inverted phase contrast microscopy for the percentage of growth cones showing a collapsed profile with loss of flattened morphology, filopodia, and lamellipodia.

  1. Radon assay for SNO+

    NASA Astrophysics Data System (ADS)

    Rumleskie, Janet

    2015-12-01

    The SNO+ experiment will study neutrinos while located 6,800 feet below the surface of the earth at SNOLAB. Though shielded from surface backgrounds, emanation of radon radioisotopes from the surrounding rock leads to back-grounds. The characteristic decay of radon and its daughters allows for an alpha detection technique to count the amount of Rn-222 atoms collected. Traps can collect Rn-222 from various positions and materials, including an assay skid that will collect Rn-222 from the organic liquid scintillator used to detect interactions within SNO+.

  2. RAS - Screens & Assays - Drug Discovery

    Cancer.gov

    The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

  3. Biosensors: Viruses for ultrasensitive assays

    NASA Astrophysics Data System (ADS)

    Donath, Edwin

    2009-04-01

    A three-dimensional assay based on genetically engineered viral nanoparticles and nickel nanohairs can detect much lower levels of protein markers associated with heart attacks than conventional assays.

  4. TOTAL CULTURABLE VIRUS QUANTAL ASSAY

    EPA Science Inventory

    This chapter describes a quantal method for assaying culturable human enteric viruses from water matrices. The assay differs from the plaque assay described in Chapter 10 (December 1987 Revision) in that it is based upon the direct microscopic viewing of cells for virus-induced ...

  5. Chemotaxis: Under Agarose Assay.

    PubMed

    Brazill, Derrick

    2016-01-01

    The unicellular eukaryote Dictyostelium discoideum represents a superb model for examining chemotaxis. Under vegetative conditions, the amoebae are chemotactically responsive to pterins, such as folate. Under starved conditions, they lose their sensitivity to pterins, and become chemotactically responsive to cAMP. As an NIH model system, Dictyostelium offers a variety of advantages in studying chemotaxis, including its conservation of mammalian signaling pathways, its ease of growth, and its genetic tractability. In this chapter, we describe the use of the under agarose chemotaxis assay to identify proteins involved in controlling motility and directional sensing in Dictyostelium discoideum. Given the similarities between Dictyostelium and mammalian cells, this allows us to dissect the conserved pathways involved in eukaryotic chemotaxis.

  6. Adipose tissue angiogenesis assay.

    PubMed

    Rojas-Rodriguez, Raziel; Gealekman, Olga; Kruse, Maxwell E; Rosenthal, Brittany; Rao, Kishore; Min, Soyun; Bellve, Karl D; Lifshitz, Lawrence M; Corvera, Silvia

    2014-01-01

    Changes in adipose tissue mass must be accompanied by parallel changes in microcirculation. Investigating the mechanisms that regulate adipose tissue angiogenesis could lead to better understanding of adipose tissue function and reveal new potential therapeutic strategies. Angiogenesis is defined as the formation of new capillaries from existing microvessels. This process can be recapitulated in vitro, by incubation of tissue in extracellular matrix components in the presence of pro-angiogenic factors. Here, we describe a method to study angiogenesis from adipose tissue fragments obtained from mouse and human tissue. This assay can be used to define effects of diverse factors added in vitro, as well as the role of endogenously produced factors on angiogenesis. We also describe approaches to quantify angiogenic potential for the purpose of enabling comparisons between subjects, thus providing information on the role of physiological conditions of the donor on adipose tissue angiogenic potential.

  7. Yeast DEL assay detects clastogens.

    PubMed

    Kirpnick, Zhanna; Homiski, Michael; Rubitski, Elizabeth; Repnevskaya, Marina; Howlett, Niall; Aubrecht, Jiri; Schiestl, Robert H

    2005-04-01

    Chromosomal rearrangements, including DNA deletions are involved in carcinogenesis. The deletion (DEL) assay scoring for DNA deletions in the yeast Saccharomyces cerevisiae is able to detect a wide range of carcinogens. Among approximately 60 compounds of known carcinogenic activity, the DEL assay detected 86% correctly whereas the Ames Salmonella assay detected only 30% correctly [R.J. Brennan, R.H. Schiestl, Detecting carcinogens with the yeast DEL assay, Methods Mol. Biol. 262 (2004) 111-124]. Since the DEL assay is highly inducible by DNA double strand breaks, this study examined the utility of the DEL assay for detecting clastogens. Ten model compounds, with varied mechanisms of genotoxicity, were examined for their effect on the frequency of DNA deletions with the DEL assay. The compounds tested were: actinomycin D, camptothecin, methotrexate and 5-fluorodeoxyuridine, which are anticancer agents, noscapine and furosemide are therapeutics, acridine, methyl acrylate and resorcinol are industrial chemicals and diazinon is an insecticide. The in vitro micronucleus assay (IVMN) in CHO cells, a commonly used tool for detection of clastogens, was performed on the same compounds and the results of the two assays were compared. The results of our study show that there is 70% concordance in the presence of metabolic activation (rat liver S9) and 80% concordance in the absence of metabolic activation between the DEL assay and the standard in vitro micronucleus assay. The lack of cytotoxicity observed for four of the ten compounds examined indicates limited diffusion of lipophilic compounds across the yeast cell wall. Thus, the development of a more permeable yeast tester strain is expected to greatly improve concordance of the DEL assay with the IVMN assay. The yeast DEL assay is inexpensive, amenable to automation and requires less expertise to perform than the IVMN. Thus, it has a strong potential as a robust, fast and economical screen for detecting clastogens in

  8. An assay for adjuvanticity

    PubMed Central

    Dresser, D. W.

    1968-01-01

    Adult mice injected with an adequate amount of a non-immunogenic antigen progress to a specific state of immunological paralysis, unless a substance with `extrinsic' adjuvanticity is injected before the induction of paralysis is completed. Consequently incipiently paralysed mice can be used to assay substances for adjuvanticity. Conventional adjuvants such as Freund's adjuvant and pertussis possess adjuvanticity; other substances with varying degrees of adjuvanticity are listed in the tables. It has been shown that the adjuvanticity effect of an injection of pertussis lasts for only a few days, although the effect of such an injection of pertussis on phagocytosis of carbon particles does not reach a maximum until 2 weeks after the injection. The dose-effectiveness of alum precipitated (highly phagocytosable) bovine γ-globulin was greatly increased by the intraperitoneal injection of pertussis. The evidence is considered to be incompatible with increased phagocytosis being either an essential factor in the role of pertussis as a conventional adjuvant, or in the adjuvanticity effect of pertussis. PMID:4179956

  9. Click Chemistry-Mediated Nanosensors for Biochemical Assays

    PubMed Central

    Chen, Yiping; Xianyu, Yunlei; Wu, Jing; Yin, Binfeng; Jiang, Xingyu

    2016-01-01

    Click chemistry combined with functional nanoparticles have drawn increasing attention in biochemical assays because they are promising in developing biosensors with effective signal transformation/amplification and straightforward signal readout for clinical diagnostic assays. In this review, we focus on the latest advances of biochemical assays based on Cu (I)-catalyzed 1, 3-dipolar cycloaddition of azides and alkynes (CuAAC)-mediated nanosensors, as well as the functionalization of nanoprobes based on click chemistry. Nanoprobes including gold nanoparticles, quantum dots, magnetic nanoparticles and carbon nanomaterials are covered. We discuss the advantages of click chemistry-mediated nanosensors for biochemical assays, and give perspectives on the development of click chemistry-mediated approaches for clinical diagnosis and other biomedical applications. PMID:27217831

  10. Practical assay issues with the PERT/PBRT assay: a highly sensitive reverse transcriptase assay.

    PubMed

    Chang, A; Dusing, S

    2006-01-01

    Product safety testing for retroviruses can be achieved by a panel of screening assays, including electron microscopy, viral gene specific PCRs, virus propagation, and detection of reverse transciptase activity. The application of PCR-based reverse transcriptase assays (PERT) that are approximately a million-fold more sensitive than conventional nucleotide incorporation assays in the testing of biologicals is described. Use of PERT assays can be applied to three areas: (i) screening for adventitious retrovirus contamination; (ii) detecting and quantifying endogenous viral particle load and (iii) monitoring levels of infectious retrovirus generation in cell lines that contain endogenous retroviruses.

  11. From Antenna to Assay

    PubMed Central

    Moore, Evan G.; Samuel, Amanda P. S.; Raymond, Kenneth N.

    2009-01-01

    Conspectus Ligand-sensitized, luminescent lanthanide(III) complexes are of considerable importance because their unique photophysical properties (microsecond to millisecond lifetimes, characteristic and narrow emission bands, and large Stokes shifts) make them well suited as labels in fluorescence-based bioassays. The long-lived emission of lanthanide(III) cations can be temporally resolved from scattered light and background fluorescence to vastly enhance measurement sensitivity. One challenge in this field is the design of sensitizing ligands that provide highly emissive complexes with sufficient stability and aqueous solubility for practical applications. In this Account, we give an overview of some of the general properties of the trivalent lanthanides and follow with a summary of advances made in our laboratory in the development of highly luminescent Tb(III) and Eu(III) complexes for applications in biotechnology. A focus of our research has been the optimization of these compounds as potential commercial agents for use in Homogeneous Time-Resolved Fluorescence (HTRF) technology. Our approach involves developing high-stability octadentate Tb(III) and Eu(III) complexes that rely on all-oxygen donor atoms and using multi-chromophore chelates to increase molar absorptivity; earlier examples utilized a single pendant chromophore (that is, a single “antenna”). Ligands based on 2-hydroxyisophthalamide (IAM) provide exceptionally emissive Tb(III) complexes with quantum yield values up to ∼60% that are stable at the nanomolar concentrations required for commercial assays. Through synthetic modification of the IAM chromophore and time-dependent density functional theory (TD-DFT) calculations, we have developed a method to predict absorption and emission properties of these chromophores as a tool to guide ligand design. Additionally, we have investigated chiral IAM ligands that yield Tb(III) complexes possessing both high quantum yield values and strong

  12. The assay of diphtheria toxin

    PubMed Central

    Gerwing, Julia; Long, D. A.; Mussett, Marjorie V.

    1957-01-01

    A precise assay of diphtheria toxin is described, based on the linear relationship between the diameter of the skin reaction to, and logarithm of the dose of, toxin. It eliminates the need for preliminary titrations, is economical, provides information about the slope of the log-dose response lines and, therefore, of the validity of the assay, and yields limits of error of potency from the internal evidence of the assay. A study has been made of the effects of avidity, combining power, toxicity and buffering on the assay of diphtheria toxins against the International Standards for both Diphtheria Antitoxin and Schick-Test Toxin. All the toxins assayed against the standard toxin, whatever their other properties might be, gave log-dose response lines of similar slope provided that they were diluted in buffered physiological saline. The assays were therefore valid. These experiments were repeated concurrently in non-immune and in actively immunized guinea-pigs, and comparable figures for potency obtained in both groups. The result was not significantly affected by the avidity or combining power of the toxin. However, non-avid toxins gave low values in Schick units when assayed, by the Römer & Sames technique, in terms of the International Standard for Diphtheria Antitoxin. The problem of the ultimate standard and the implications of these findings are discussed. PMID:13511133

  13. Comet Assay measurements: a perspective.

    PubMed

    Kumaravel, T S; Vilhar, Barbara; Faux, Stephen P; Jha, Awadhesh N

    2009-02-01

    The Comet Assay or single cell gel electrophoresis assay is one of the very widely used assays to microscopically detect DNA damage at the level of a single cell. The determination of damage is carried out either through visual scoring of cells (after classification into different categories on the basis of tail length and shape) or by using different commercially available or public domain software (which automatically recognise the extent of damage). In this assay, the shape, size and amount of DNA within the 'comet' play important roles in the determination of the level of damage. The use of a software in particular also provides a range of different parameters, many of which might not be relevant in determining the extent of DNA damage. As a large number of factors could influence the shape, size, identification and determination of induced damage, which includes the scoring criteria, staining techniques, selection of parameters (whilst using the software packages) and appearance of 'hedgehog' or 'clouds', this article aims (a) to provide an overview of evolution of measurements of DNA damage using the Comet Assay and (b) to summarise and critically analyse the advantages and disadvantages of different approaches currently being adopted whilst using this assay. It is suggested that judicious selection of different parameters, staining methods along with inter-laboratory validation and harmonisation of methodologies will further help in making this assay more robust and widely acceptable for scientific as well as regulatory studies.

  14. Plaque assay for murine norovirus.

    PubMed

    Gonzalez-Hernandez, Mariam B; Bragazzi Cunha, Juliana; Wobus, Christiane E

    2012-01-01

    Murine norovirus (MNV) is the only member of the Norovirus genus that efficiently grows in tissue culture. Cell lysis and cytopathic effect (CPE) are observed during MNV-1 infection of murine dendritic cells or macrophages. This property of MNV-1 can be used to quantify the number of infectious particles in a given sample by performing a plaque assay. The plaque assay relies on the ability of MNV-1 to lyse cells and to form holes in a confluent cell monolayer, which are called plaques. Multiple techniques can be used to detect viral infections in tissue culture, harvested tissue, clinical, and environmental samples, but not all measure the number of infectious particles (e.g. qRT-PCR). One way to quantify infectious viral particles is to perform a plaque assay, which will be described in detail below. A variation on the MNV plaque assay is the fluorescent focus assay, where MNV antigen is immunostained in cell monolayers. This assay can be faster, since viral antigen expression precedes plaque formation. It is also useful for titrating viruses unable to form plaques. However, the fluorescent focus assay requires additional resources beyond those of the plaque assay, such as antibodies and a microscope to count focus-forming units. Infectious MNV can also be quantified by determining the 50% Tissue Culture Infective Dose (TCID50). This assay measures the amount of virus required to produce CPE in 50% of inoculated tissue culture cells by endpoint titration. However, its limit of detection is higher compared to a plaque assay. In this article, we describe a plaque assay protocol that can be used to effectively determine the number of infectious MNV particles present in biological or environmental samples. This method is based on the preparation of 10-fold serial dilutions of MNV-containing samples, which are used to inoculate a monolayer of permissive cells (RAW 264.7 murine macrophage cells). Virus is allowed to attach to the cell monolayer for a given period of

  15. Troponin revisited 2008: assay performance.

    PubMed

    Tate, Jillian R

    2008-01-01

    Troponin quality specifications describing the pre-analytical, analytical and post-analytical performance of cardiac troponin (cTn) assays are important for both manufacturers of cTn assays and laboratories that routinely test for cTn. Pre-analytical requirements refer not only to acceptable sample type for analysis and the stability of cTn but also to the proper handling of specimens prior to analysis to avoid pre-analytical false positive results. Analytical issues that may contribute to differences between cTn assays include analytical sensitivity and imprecision at low cTn concentration, antibody specificity and immunoreactivity of plasma cTn forms, assay specificity and the presence of falsely positive and negative interferences, and for cTnI the lack of standardised measurement, all which may impact on patient cTn results. Current second generation cTnI and fourth generation cTnT assays generally have an imprecision of around 20% coefficient of variation (CV) at the 99th percentile of the reference population, which is greater than the recommended imprecision of 10% CV. As the next generation of more analytically sensitive cTn assays are developed it can be anticipated that cTn upper reference limits will decrease by approximately 10-fold. Monitoring assay imprecision at ultra low cTn concentrations will require that the laboratory uses a quality control close to this level and a negative control to monitor baseline drift. Establishment of cTn reference ranges will require reference populations to be cardio-healthy to enable differentiation from community populations who are at increased cardiovascular risk. Close collaboration between the laboratory and local clinicians is required to ensure adequate clinical validation of more sensitive cTn assays.

  16. Methods to assay Drosophila behavior.

    PubMed

    Nichols, Charles D; Becnel, Jaime; Pandey, Udai B

    2012-01-01

    Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases(1). We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials(2-4). The rapid iterative negative geotaxis (RING) assay(5) has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously

  17. Microbiological assay using bioluminescent organism

    SciTech Connect

    Stiffey, A.V.

    1987-12-21

    This invention relates to testing processes for toxicity involving microorganisms and, more particularly, to testing processes for toxicity involving bioluminescent organisms. The present known method of testing oil-well drilling fluids for toxicity employs the mysid shrimp (Mysidopsis bahia) as the assay organism. The shrimp are difficult to raise and handle as laboratory assay organisms. This method is labor-intensive, because it requires a assay time of about 96 hours. Summary of the Invention: A microbiological assay in which the assay organism is the dinoflagellate, Pyrocystis lunula. A sample of a substance to be assayed is added to known numbers of the bioluminescent dinoflagellate and the mixture is agitated to subject the organisms to a shear stress causing them to emit light. The amount of light emitted is measured and compared with the amount of light emitted by a known non-toxic control mixture to determine if there is diminution or non-diminution of light emitted by the sample under test which is an indication of the presence or absence of toxicity, respectively. Accordingly, an object of the present invention is the provision of an improved method of testing substances for toxicity. A further object of the invention is the provision of an improved method of testing oil-well drilling fluids for toxicity using bioluminescent dinoflagellate (Pyrocystis lunula).

  18. Assaying DNA damage in hippocampal neurons using the comet assay.

    PubMed

    Nowsheen, Somaira; Xia, Fen; Yang, Eddy S

    2012-12-19

    A number of drugs target the DNA repair pathways and induce cell kill by creating DNA damage. Thus, processes to directly measure DNA damage have been extensively evaluated. Traditional methods are time consuming, expensive, resource intensive and require replicating cells. In contrast, the comet assay, a single cell gel electrophoresis assay, is a faster, non-invasive, inexpensive, direct and sensitive measure of DNA damage and repair. All forms of DNA damage as well as DNA repair can be visualized at the single cell level using this powerful technique. The principle underlying the comet assay is that intact DNA is highly ordered whereas DNA damage disrupts this organization. The damaged DNA seeps into the agarose matrix and when subjected to an electric field, the negatively charged DNA migrates towards the cathode which is positively charged. The large undamaged DNA strands are not able to migrate far from the nucleus. DNA damage creates smaller DNA fragments which travel farther than the intact DNA. Comet Assay, an image analysis software, measures and compares the overall fluorescent intensity of the DNA in the nucleus with DNA that has migrated out of the nucleus. Fluorescent signal from the migrated DNA is proportional to DNA damage. Longer brighter DNA tail signifies increased DNA damage. Some of the parameters that are measured are tail moment which is a measure of both the amount of DNA and distribution of DNA in the tail, tail length and percentage of DNA in the tail. This assay allows to measure DNA repair as well since resolution of DNA damage signifies repair has taken place. The limit of sensitivity is approximately 50 strand breaks per diploid mammalian cell (1,2). Cells treated with any DNA damaging agents, such as etoposide, may be used as a positive control. Thus the comet assay is a quick and effective procedure to measure DNA damage.

  19. Magnetic Spinner

    NASA Astrophysics Data System (ADS)

    Ouseph, P. J.

    2006-12-01

    A science toy sometimes called the "magnetic spinner" is an interesting class demonstration to illustrate the principles of magnetic levitation. It can also be used to demonstrate Faraday's law and a horizontally suspended physical pendulum. The levitated part contains two circular magnets encased in a plastic housing. Each magnet stays above two triangular magnets fixed to the base. The magnetic repulsive force experienced by the circular magnets is independent of their orientation; therefore, the holder of these magnets can be rotated without affecting its stability. The holder with the circular magnets can be oscillated up and down as a horizontally suspended physical pendulum.

  20. Immunomagnetic nanoparticle-based assays for detection of biomarkers

    PubMed Central

    Park, Hoyoung; Hwang, Mintai P; Lee, Kwan Hyi

    2013-01-01

    The emergence of biomarkers as key players in the paradigm shift towards preventative medicine underscores the need for their detection and quantification. Advances made in the field of nanotechnology have played a crucial role in achieving these needs, and have contributed to recent advances in the field of medicine. Nanoparticle-based immunomagnetic assays, in particular, offer numerous advantages that utilize the unique physical properties of magnetic nanoparticles. In this review, we focus on recent developments and trends with regards to immunomagnetic assays used for detection of biomarkers. The various immunomagnetic assays are categorized into the following: particle-based multiplexing, signal control, microfluidics, microarray, and automation. Herein, we analyze each category and discuss their advantages and disadvantages. PMID:24285924

  1. Optimization of protein samples for NMR using thermal shift assays.

    PubMed

    Kozak, Sandra; Lercher, Lukas; Karanth, Megha N; Meijers, Rob; Carlomagno, Teresa; Boivin, Stephane

    2016-04-01

    Maintaining a stable fold for recombinant proteins is challenging, especially when working with highly purified and concentrated samples at temperatures >20 °C. Therefore, it is worthwhile to screen for different buffer components that can stabilize protein samples. Thermal shift assays or ThermoFluor(®) provide a high-throughput screening method to assess the thermal stability of a sample under several conditions simultaneously. Here, we describe a thermal shift assay that is designed to optimize conditions for nuclear magnetic resonance studies, which typically require stable samples at high concentration and ambient (or higher) temperature. We demonstrate that for two challenging proteins, the multicomponent screen helped to identify ingredients that increased protein stability, leading to clear improvements in the quality of the spectra. Thermal shift assays provide an economic and time-efficient method to find optimal conditions for NMR structural studies. PMID:26984476

  2. HIV-1 Capsid Stabilization Assay.

    PubMed

    Fricke, Thomas; Diaz-Griffero, Felipe

    2016-01-01

    The stability of the HIV-1 core in the cytoplasm is crucial for productive HIV-1 infection. Mutations that stabilize or destabilize the core showed defects in HIV-1 reverse transcription and infection. We developed a novel and simple assay to measure stability of in vitro-assembled HIV-1 CA-NC complexes. This assay allowed us to demonstrate that cytosolic extracts strongly stabilize the HIV-1 core (Fricke et al., J Virol 87:10587-10597, 2013). By using our novel assay, one can measure the ability of different drugs to modulate the stability of in vitro-assembled HIV-1 CA-NC complexes, such as PF74, CAP-1, IXN-053, cyclosporine A, Bi2, and the peptide CAI. We also found that purified CPSF6 (1-321) protein stabilizes in vitro-assembled HIV-1 CA-NC complexes (Fricke et al., J Virol 87:10587-10597, 2013). Here we describe in detail the use of this capsid stability assay. We believe that our assay can be a powerful tool to assess HIV-1 capsid stability in vitro.

  3. Time-Resolved Fluorescence Assays.

    PubMed

    Ma, Chen-Ting; Sergienko, Eduard A

    2016-01-01

    Fluorescence-based detection techniques are popular in high throughput screening due to sensitivity and cost-effectiveness. Four commonly used techniques exist, each with distinct characteristics. Fluorescence intensity assays are the simplest to run, but suffer the most from signal interference. Fluorescence polarization assays show less interference from the compounds or the instrument, but require a design that results in change of fluorophore-containing moiety size and usually have narrow assay signal window. Fluorescence resonance energy transfer (FRET) is commonly used for detecting protein-protein interactions and is constrained not by the sizes of binding partners, but rather by the distance between fluorophores. Time-resolved fluorescence resonance energy transfer (TR-FRET), an advanced modification of FRET approach utilizes special fluorophores with long-lived fluorescence and earns its place near the top of fluorescent techniques list by its performance and robustness, characterized by larger assay window and minimized compound spectral interference. TR-FRET technology can be applied in biochemical or cell-based in vitro assays with ease. It is commonly used to detect modulation of protein-protein interactions and in detection of products of biochemical reactions and cellular activities. PMID:27316992

  4. Luminogenic cytochrome P450 assays.

    PubMed

    Cali, James J; Ma, Dongping; Sobol, Mary; Simpson, Daniel J; Frackman, Susan; Good, Troy D; Daily, William J; Liu, David

    2006-08-01

    Luminogenic cytochrome P450 (CYP) assays couple CYP enzyme activity to firefly luciferase luminescence in a technology called P450-Glo(TM) (Promega). Luminogenic substrates are used in assays of human CYP1A1, -1A2, -1B1, -2C8, -2C9, -2C19, -2D6, -2J2, -3A4, -3A7, -4A11, -4F3B, -4F12 and -19. The assays detect dose-dependent CYP inhibition by test compounds against recombinant CYP enzymes or liver microsomes. Induction or inhibition of CYP activities in cultured hepatocytes is measured in a nonlytic approach that leaves cells intact for additional analysis. Luminogenic CYP assays offer advantages of speed and safety over HPLC and radiochemical-based methods. Compared with fluorogenic methods the approach offers advantages of improved sensitivity and decreased interference between optical properties of test compound and CYP substrate. These homogenous assays are sensitive and robust tools for high-throughput CYP screening in early drug discovery. PMID:16859410

  5. Barcoded microchips for biomolecular assays.

    PubMed

    Zhang, Yi; Sun, Jiashu; Zou, Yu; Chen, Wenwen; Zhang, Wei; Xi, Jianzhong Jeff; Jiang, Xingyu

    2015-01-20

    Multiplexed assay of analytes is of great importance for clinical diagnostics and other analytical applications. Barcode-based bioassays with the ability to encode and decode may realize this goal in a straightforward and consistent manner. We present here a microfluidic barcoded chip containing several sets of microchannels with different widths, imitating the commonly used barcode. A single barcoded microchip can carry out tens of individual protein/nucleic acid assays (encode) and immediately yield all assay results by a portable barcode reader or a smartphone (decode). The applicability of a barcoded microchip is demonstrated by human immunodeficiency virus (HIV) immunoassays for simultaneous detection of three targets (anti-gp41 antibody, anti-gp120 antibody, and anti-gp36 antibody) from six human serum samples. We can also determine seven pathogen-specific oligonucleotides by a single chip containing both positive and negative controls.

  6. Displacement enzyme linked aptamer assay.

    PubMed

    Baldrich, Eva; Acero, Josep Lluis; Reekmans, Gunter; Laureyn, Wim; O'Sullivan, Ciara K

    2005-08-01

    Immense effort has been placed on the realization of immunoassays exploiting displacement of a suboptimum target, due to the ease of use and applicability to immunochromatographic strips and immunosensors. Most of the efforts reported to date focus on the use of a suboptimal target that is displaceable by the target toward which the antibody has higher affinity. Limited success has been achieved due to difficulty in obtaining suboptimal targets to which the antibody has enough affinity to bind while at the same time having lower levels of affinity in comparison to the target to facilitate displacement. Aptamers are synthetic oligonucleotides specifically selected to bind a certain target. Thanks to their high affinity and sensitivity, aptamers appear as alternative candidates to antibodies for analytical devices and several enzyme-linked aptamer assays and aptasensors have been reported. Aptamers, in contrast to antibodies, require the formation of a three-dimensional structure for target binding and can thus be anticipated to have a much higher affinity for binding its target rather than a modified form of the target (e.g., enzyme-labeled target). This phenomenon can be exploited for the development of a displacement assay, using enzyme-labeled target as a suboptimal displaceable molecule. Here, we report the first demonstration of the exploitation of an aptamer in an extremely rapid and highly sensitive displacement assay. Surface plasmon resonance studies demonstrated the thrombin-binding aptamer to have a lower affinity for enzyme-labeled thrombin than unmodified thrombin, with respective K(D) of 1.1 x 10(-8) and 2.9 x 10(-9) M. The assay is extremely rapid, requiring only 10 min for completion, and exhibits a detection limit lower than that obtainable with competitive enzyme-linked aptamer assays and comparable to that of hybrid aptamer-antibody assays. Optimal storage conditions for precoated microtiter plates (consisting of coated aptamer and captured

  7. Neodymium Magnets.

    ERIC Educational Resources Information Center

    Wida, Sam

    1992-01-01

    Uses extremely strong neodymium magnets to demonstrate several principles of physics including electromagnetic induction, Lenz's Law, domain theory, demagnetization, the Curie point, and magnetic flux lines. (MDH)

  8. Three dimensional colorimetric assay assemblies

    SciTech Connect

    Charych, D.; Reichart, A.

    2000-06-27

    A direct assay is described using novel three-dimensional polymeric assemblies which change from a blue to red color when exposed to an analyte, in one case a flu virus. The assemblies are typically in the form of liposomes which can be maintained in a suspension, and show great intensity in their color changes. Their method of production is also described.

  9. Assays for B lymphocyte function.

    PubMed

    Bondada, Subbarao; Robertson, Darrell A

    2003-11-01

    This unit describes the antigenic stimulation of in vitro antibody production by B cells and the subsequent measurement of secreted antibodies. The first basic protocol is a generalized system for inducing in vitro antibody production and can accommodate various types of antigens under study. Secreted antibodies can then be measured with an enzyme-linked immunosorbent assay (ELISA) or other soluble-antibody detection systems. Alternatively, the number of antibody-producing cells can be quantified by plaque-forming cell (PFC) assays presented in this unit: the Cunningham-Szenberg and the Jerne-Nordin techniques. Both methods employ specially prepared slide chambers, described here, in which the antibody-producing B cells are mixed with complement and indicator sheep red blood cells (SRBC), or with trinitrophenol-modified SRBC (TNP-SRBC), with subsequent lysis and counting of plaques. Because IgM antibodies fix complement efficiently, whereas IgG and IgA antibodies do not, unmodified PFC assays measure only IgM antibodies. The assay can be modified, however, to measure all classes of antibodies or to enumerate total immunoglobulin-secreting B cells, as described in alternate protocols. Yet another method of measuring the number of antibody-producing B cells (in a class-specific fashion) is to use the ELISPOT technique described in UNIT 7.14. The resting B cells used in these procedures are prepared as described in the final support protocols for Percoll gradient centrifugation. PMID:18432909

  10. Bacterial mutagenicity assays: test methods.

    PubMed

    Gatehouse, David

    2012-01-01

    The most widely used assays for detecting chemically induced gene mutations are those employing bacteria. The plate incorporation assay using various Salmonella typhimurium LT2 and E. coli WP2 strains is a short-term bacterial reverse mutation assay specifically designed to detect a wide range of chemical substances capable of causing DNA damage leading to gene mutations. The test is used worldwide as an initial screen to determine the mutagenic potential of new chemicals and drugs.The test uses several strains of S. typhimurium which carry different mutations in various genes of the histidine operon, and E. coli which carry the same AT base pair at the critical mutation site within the trpE gene. These mutations act as hot spots for mutagens that cause DNA damage via different mechanisms. When these auxotrophic bacterial strains are grown on a minimal media agar plates containing a trace of the required amino-acid (histidine or tryptophan), only those bacteria that revert to amino-acid independence (His(+) or Tryp(+)) will grow to form visible colonies. The number of spontaneously induced revertant colonies per plate is relatively constant. However, when a mutagen is added to the plate, the number of revertant colonies per plate is increased, usually in a dose-related manner.This chapter provides detailed procedures for performing the test in the presence and absence of a metabolic activation system (S9-mix), including advice on specific assay variations and any technical problems. PMID:22147566

  11. An improved choline monooxygenase assay

    SciTech Connect

    Lafontaine, P.J.; Hanson, A.D. )

    1991-05-01

    Glycine betaine accumulates in leaves of plants from several angiosperm families in response to drought or salinization. Its synthesis, from the oxidation of choline, is mediated by a two step pathway. In spinach the first enzyme of this pathway is a ferredoxin-dependent choline monooxygenase (CMO). In order to purify this enzyme a sensitive and reliable assay is necessary. Two types of modifications were explored to improve the existing assay. (1) Ferredoxin reduction - one way of providing reduced Fd to CMO is by the addition of isolated spinach thylakoids in the assay mixture. In order to optimize the reduction of Fd two different systems were compared: (a) where only PS is active, by adding DCMU to inhibit electron transport from PS II and DAD as electron donor for PS I; (b) where both PS II and PS I are active. (2) Betaine aldehyde estimation - to simplify this, it is possible to couple the CMO reaction with betaine aldehyde dehydrogenase (BADH) from E. coli. BADH converts betaine aldehyde to betaine as it is formed in the assay, eliminating the need for a chemical oxidation step.

  12. Three dimensional colorimetric assay assemblies

    DOEpatents

    Charych, Deborah; Reichart, Anke

    2000-01-01

    A direct assay is described using novel three-dimensional polymeric assemblies which change from a blue to red color when exposed to an analyte, in one case a flu virus. The assemblies are typically in the form of liposomes which can be maintained in a suspension, and show great intensity in their color changes. Their method of production is also described.

  13. Recent Developments in Electrotaxis Assays.

    PubMed

    Wu, Jiandong; Lin, Francis

    2014-02-01

    Significance: A wide range of cell types can migrate in response to physiological or externally applied direct current electric field (dcEF), a process termed electrotaxis. In particular, electrotaxis of epithelial cells to wound-generated dcEF for mediating wound healing is a well-accepted mechanism. In addition, various immune cells have been demonstrated to undergo electrotaxis, suggesting a link between electrotaxis and inflammatory responses in wound healing. Electrotaxis research will generate important insight into the electrical guiding mechanism for cell migration thereby providing the scientific basis to further develop clinical applications for wound care. Development of advanced electrotaxis assays will critically enable in-depth experimental electrotaxis studies in vitro. Recent Advances: Recently, a number of new electrotaxis assays or new uses of previously developed assays for electrotaxis studies have been reported. These new developments provide improved solutions for experimental throughput, configuration of three-dimensional cell migration environments and coexisting guiding signals, measurements of collective electrotactic cell migration, and sorting electrotactic populations. Critical Issues: These new developments face the challenge of playing a more important role to better understand the biological mechanisms underlying electrotaxis, in addition to making a stronger impact on relevant applications. Future Directions: On one hand, specific electrotaxis assays should be further developed to improve its function and tested for a broader range of experimental conditions and electrotactic populations. On the other hand, joint efforts among electrotaxis researchers are needed to integrate the unique features of specific electrotaxis assays, allowing more advanced and efficient electrotaxis analyses to answer both basic science and clinical questions. PMID:24761355

  14. Magnetic Relaxation Detector for Microbead Labels

    PubMed Central

    Liu, Paul Peng; Skucha, Karl; Duan, Yida; Megens, Mischa; Kim, Jungkyu; Izyumin, Igor I.; Gambini, Simone; Boser, Bernhard

    2014-01-01

    A compact and robust magnetic label detector for biomedical assays is implemented in 0.18-μm CMOS. Detection relies on the magnetic relaxation signature of a microbead label for improved tolerance to environmental variations and relaxed dynamic range requirement, eliminating the need for baseline calibration and reference sensors. The device includes embedded electromagnets to eliminate external magnets and reduce power dissipation. Correlated double sampling combined with offset servo loops and magnetic field modulation, suppresses the detector offset to sub-μT. Single 4.5-μm magnetic beads are detected in 16 ms with a probability of error <0.1%. PMID:25308988

  15. Broad base biological assay using liquid based detection assays

    SciTech Connect

    Milanovich, F; Albala, J; Colston, B; Langlois, R; Venkateswaren, K

    2000-10-31

    The release of a biological agent by terrorists represents a serious threat to the safety of US citizens. At present there are over 50 pathogens and toxins on various agency threat lists. Most of these pathogens are rarely seen by public health personnel so the ability to rapidly identify their infection is limited. Since many pathogenic infections have symptomatic delays as long as several days, effective treatment is often compromised. This translates into two major deficiencies in our ability to counter biological terrorism (1) the lack of any credible technology to rapidly detect and identify all the pathogens or toxins on current threat lists and (2) the lack of a credible means to rapidly diagnose thousands of potential victims. In this SI we are developing a rapid, flexible, inexpensive, high throughput, and deeply multiplex-capable biological assay technology. The technology, which we call the Liquid Array (LA), utilizes optical encoding of small diameter beads which serve as the templates for biological capture assays. Once exposed to a fluid sample these beads can be identified and probed for target pathogens at rates of several thousand beads per second. Since each bead can be separately identified, one can perform parallel assays by assigning a different assay to each bead in the encoded set. The goal for this development is a detection technology capable of simultaneously identifying 100s of different bioagents and/or of rapidly diagnosing several thousand individuals. We are pursuing this research in three thrusts. In the first we are exploring the fundamental interactions of the beads with proteins and nucleic acids in complex mixtures. This will provide us with a complete understanding of the limits of the technology with respect to throughput and complex environment. A major spin-off of this activity is in the rapidly emerging field of proteomics where we may be able to rapidly assess the interactions responsible for cell metabolism, structural

  16. 21 CFR 225.158 - Laboratory assays.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 4 2012-04-01 2012-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  17. 21 CFR 225.158 - Laboratory assays.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  18. 21 CFR 225.158 - Laboratory assays.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 4 2014-04-01 2014-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  19. 21 CFR 225.158 - Laboratory assays.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 4 2011-04-01 2011-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  20. 21 CFR 225.158 - Laboratory assays.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 4 2013-04-01 2013-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  1. Magnetic Spinner

    ERIC Educational Resources Information Center

    Ouseph, P. J.

    2006-01-01

    A science toy sometimes called the "magnetic spinner" is an interesting class demonstration to illustrate the principles of magnetic levitation. It can also be used to demonstrate Faraday's law and a horizontally suspended physical pendulum. The levitated part contains two circular magnets encased in a plastic housing. Each magnet stays…

  2. Magnetic Levitation.

    ERIC Educational Resources Information Center

    Rossing, Thomas D.; Hull, John R.

    1991-01-01

    Discusses the principles of magnetic levitation presented in the physics classroom and applied to transportation systems. Topics discussed include three classroom demonstrations to illustrate magnetic levitation, the concept of eddy currents, lift and drag forces on a moving magnet, magnetic levitation vehicles, levitation with permanent magnets…

  3. Important Norwegian crude assays updated

    SciTech Connect

    Corbett, R.A

    1990-03-12

    New assays on two important Norwegian North Sea crude oils, Statfjord and Gullfaks, are presented. Both are high-quality, low-sulfur crudes that will yield a full range of good-quality products. All assay data came from industry-standard test procedures. The Statfjord field is the largest in the North Sea. Production started in 1979. Statfjord is a typical North Sea crude, produced from three separate platforms and three separate loading buoys with interconnecting lines. Current production is about 700,000 b/d. Gullfaks is produced from a large field in Block 34/10 of the Norwegian sector of the North Sea production area. Gullfaks crude oil is more biodegraded than other crudes from the region. Biodegradation has removed most of the waxy normal paraffins, resulting in a heavier, more naphthenic and aromatic crude.

  4. The skin-blanching assay.

    PubMed

    Smit, P; Neumann, H A M; Thio, H B

    2012-10-01

    The skin-blanching assay is used for the determination and bioequivalence of dermatologic glucocorticoids (GCs). The exact mechanism of the production of blanching is not fully understood, but it is considered that local vasoconstriction of the skin microvasculature and the consequent blood-flow reduction cause this phenomenon. Several factors influence skin blanching, including drug concentration, duration of application, nature of vehicle, occlusion, posture and location. The intensity of vasoconstriction can be measured in several ways: visual or quantitative methods, such as reflectance spectroscopy, thermography, laser Doppler velocimetry and chromametry. In literature, contradicting results in the correlation of the skin-blanching assay with different tests to determine GC sensitivity have been reported, limiting its clinical usefulness.

  5. Pulsating bead-based assay.

    PubMed

    Thompson, Jason A; Bau, Haim H

    2011-04-15

    In recent years, there has been a growing interest in using porous microbeads such as agarose beads as solid supports to bind target molecules from complex fluid samples. Porous beads have large surface area to volume ratios and high receptor concentrations, and they facilitate relatively high sensitivity detection and multiplexing. Unfortunately, to take full advantage of the porous beads' attributes, long incubation times are needed due to the relatively slow mass transfer of target molecules from the exterior solution into the beads' interior. To accelerate the mass transfer process, we propose a novel assay in which functionalized porous beads are periodically compressed and expanded. Preliminary experiments were carried out to compare the performance of the pulsating beads with that of conventional, nonpulsating beads. These experiments indicate that the pulsating beads significantly accelerate binding rates with minimal increase in nonspecific binding. Thus, pulsing has the potential of significantly reducing assay time.

  6. Comet Assay in Cancer Chemoprevention.

    PubMed

    Santoro, Raffaela; Ferraiuolo, Maria; Morgano, Gian Paolo; Muti, Paola; Strano, Sabrina

    2016-01-01

    The comet assay can be useful in monitoring DNA damage in single cells caused by exposure to genotoxic agents, such as those causing air, water, and soil pollution (e.g., pesticides, dioxins, electromagnetic fields) and chemo- and radiotherapy in cancer patients, or in the assessment of genoprotective effects of chemopreventive molecules. Therefore, it has particular importance in the fields of pharmacology and toxicology, and in both environmental and human biomonitoring. It allows the detection of single strand breaks as well as double-strand breaks and can be used in both normal and cancer cells. Here we describe the alkali method for comet assay, which allows to detect both single- and double-strand DNA breaks.

  7. Danish North Sea crude assayed

    SciTech Connect

    Rhodes, A.K.

    1994-09-12

    Danish North Sea blend was assayed earlier this year. The light, sweet crude comprises crude oil from 10 fields. The crude is piped from offshore production facilities to the A/S Dansk Shell refinery at Fredericia, Denmark. Fig. 1 shows the boiling point curve for the crude, and Fig. 2 illustrates the metals content (vanadium, nickel, and iron), as a function of distillation temperature. The table lists properties of the crude and its fractions.

  8. Two offshore Australian crudes assayed

    SciTech Connect

    Rhodes, A.K.

    1994-05-09

    Two light, sweet crudes from offshore Australia have been assayed. Gippsland crude, also called Bass Strait, is produced off the coast of Victoria, in southeastern Australia. The 47 API, 0.09% sulfur crude was analyzed in mid-1993. Skua, a 42 API, 0.06 wt % sulfur crude, is produced in the Timor Sea. Data are given on the whole crude and fractions for both deposits. Both chemical and physical properties are listed.

  9. Recombinase Polymerase Amplification Assay for Rapid Diagnostics of Dengue Infection

    PubMed Central

    Abd El Wahed, Ahmed; Patel, Pranav; Faye, Oumar; Thaloengsok, Sasikanya; Heidenreich, Doris; Matangkasombut, Ponpan; Manopwisedjaroen, Khajohnpong; Sakuntabhai, Anavaj; Sall, Amadou A.; Hufert, Frank T.; Weidmann, Manfred

    2015-01-01

    Background Over 2.5 billion people are exposed to the risk of contracting dengue fever (DF). Early diagnosis of DF helps to diminish its burden on public health. Real-time reverse transcription polymerase amplification assays (RT-PCR) are the standard method for molecular detection of the dengue virus (DENV). Real-time RT-PCR analysis is not suitable for on-site screening since mobile devices are large, expensive, and complex. In this study, two RT-recombinase polymerase amplification (RT-RPA) assays were developed to detect DENV1-4. Methodology/Principal Findings Using two quantitative RNA molecular standards, the analytical sensitivity of a RT-RPA targeting the 3´non-translated region of DENV1-4 was found to range from 14 (DENV4) to 241 (DENV1-3) RNA molecules detected. The assay was specific and did not cross detect other Flaviviruses. The RT-RPA assay was tested in a mobile laboratory combining magnetic-bead based total nucleic acid extraction and a portable detection device in Kedougou (Senegal) and in Bangkok (Thailand). In Kedougou, the RT-RPA was operated at an ambient temperature of 38°C with auxiliary electricity tapped from a motor vehicle and yielded a clinical sensitivity and specificity of 98% (n=31) and 100% (n=23), respectively. While in the field trial in Bangkok, the clinical sensitivity and specificity were 72% (n=90) and 100%(n=41), respectively. Conclusions/Significance During the first 5 days of infection, the developed DENV1-4 RT-RPA assays constitute a suitable accurate and rapid assay for DENV diagnosis. Moreover, the use of a portable fluorescence-reading device broadens its application potential to the point-of-care for outbreak investigations. PMID:26075598

  10. The in vitro micronucleus assay.

    PubMed

    Doherty, Ann T

    2012-01-01

    The in vitro micronucleus test detects genotoxic damage in interphase cells. The in vitro micronucleus test provides an alterative to the chromosome aberration test, and because the in vitro micronucleus test examines cells at interphase, the assessment of micronuclei can be scored faster, as the analysis of damage is thought to be less subjective and is more amenable to automation.Micronuclei may be the result of aneugenic (whole chromosome) or clastogenic (chromosome breakage) damage. This chapter provides methods for mononucleate and binucleate micronucleus tests and the addition of centromeric labelling and a non-disjunction assay to investigate any potential aneugenic mode of action.

  11. Planetary magnetism

    NASA Technical Reports Server (NTRS)

    Russell, C. T.

    1981-01-01

    A synoptic view of early and recent data on the planetary magnetism of Mercury, Venus, the moon, Mars, Jupiter, and Saturn is presented. The data on Mercury from Mariner 10 are synthesized with various other sources, while data for Venus obtained from 120 orbits of Pioneer Venus give the upper limit of the magnetic dipole. Explorer 35 Lunar Orbiter data provided the first evidence of lunar magnetization, but it was the Apollo subsatellite data that measured accurately the magnetic dipole of the moon. A complete magnetic survey of Mars is still needed, and only some preliminary data are given on the magnetic dipole of the planet. Figures on the magnetic dipoles of Jupiter and Saturn are also suggested. It is concluded that if the magnetic field data are to be used to infer the interior properties of the planets, good measures of the multiple harmonics in the field are needed, which may be obtained only through low altitude polar orbits.

  12. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  13. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  14. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  15. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  16. 21 CFR 864.7250 - Erythropoietin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... assay. (a) Identification. A erythropoietin assay is a device that measures the concentration of erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This...

  17. TRENDS IN BOREHOLE GEOPHYSICS FOR MINERAL EXPLORATION: ASSAYING AND REMOTE DETECTION.

    USGS Publications Warehouse

    Daniels, Jeffrey J.

    1985-01-01

    Several borehole geophysical techniques have been developed in recent years. Assaying technique development has been concentrated on nuclear methods, with some progress being made on using electrical and magnetic properties for mineral identification. Adaptation of conventional surface geophysical techniques to the borehole for locating near-misses of mineralized zones has led to the development of borehole resistivity, electromagnetic (EM), gravity and magnetic methods to the borehole environment. This paper discusses some of the applications and pitfalls of these new techniques.

  18. SNO+ Scintillator Purification and Assay

    NASA Astrophysics Data System (ADS)

    Ford, R.; Chen, M.; Chkvorets, O.; Hallman, D.; Vázquez-Jáuregui, E.

    2011-04-01

    We describe the R&D on the scintillator purification and assay methods and technology for the SNO+ neutrino and double-beta decay experiment. The SNO+ experiment is a replacement of the SNO heavy water with liquid scintillator comprised of 2 g/L PPO in linear alkylbenzene (LAB). During filling the LAB will be transported underground by rail car and purified by multi-stage distillation and steam stripping at a flow rate of 19 LPM. While the detector is operational the scintillator can be recirculated at 150 LPM (full detector volume in 4 days) to provide repurification as necessary by either water extraction (for Ra, K, Bi) or by functional metal scavenger columns (for Pb, Ra, Bi, Ac, Th) followed by steam stripping to remove noble gases and oxygen (Rn, O2, Kr, Ar). The metal scavenger columns also provide a method for scintillator assay for ex-situ measurement of the U and Th chain radioactivity. We have developed "natural" radioactive spikes of Pb and Ra in LAB and use these for purification testing. Lastly, we present the planned operating modes and purification strategies and the plant specifications and design.

  19. SNO+ Scintillator Purification and Assay

    SciTech Connect

    Ford, R.; Vazquez-Jauregui, E.; Chen, M.; Chkvorets, O.; Hallman, D.

    2011-04-27

    We describe the R and D on the scintillator purification and assay methods and technology for the SNO+ neutrino and double-beta decay experiment. The SNO+ experiment is a replacement of the SNO heavy water with liquid scintillator comprised of 2 g/L PPO in linear alkylbenzene (LAB). During filling the LAB will be transported underground by rail car and purified by multi-stage distillation and steam stripping at a flow rate of 19 LPM. While the detector is operational the scintillator can be recirculated at 150 LPM (full detector volume in 4 days) to provide repurification as necessary by either water extraction (for Ra, K, Bi) or by functional metal scavenger columns (for Pb, Ra, Bi, Ac, Th) followed by steam stripping to remove noble gases and oxygen (Rn, O{sub 2}, Kr, Ar). The metal scavenger columns also provide a method for scintillator assay for ex-situ measurement of the U and Th chain radioactivity. We have developed ''natural'' radioactive spikes of Pb and Ra in LAB and use these for purification testing. Lastly, we present the planned operating modes and purification strategies and the plant specifications and design.

  20. Serum indices: managing assay interference.

    PubMed

    Farrell, Christopher-John L; Carter, Andrew C

    2016-09-01

    Clinical laboratories frequently encounter samples showing significant haemolysis, icterus or lipaemia. Technical advances, utilizing spectrophotometric measurements on automated chemistry analysers, allow rapid and accurate identification of such samples. However, accurate quantification of haemolysis, icterus and lipaemia interference is of limited value if laboratories do not set rational alert limits, based on sound interference testing experiments. Furthermore, in the context of increasing consolidation of laboratories and the formation of laboratory networks, there is an increasing requirement for harmonization of the handling of haemolysis, icterus and lipaemia-affected samples across different analytical platforms. Harmonization may be best achieved by considering both the analytical aspects of index measurement and the possible variations in the effects of haemolysis, icterus and lipaemia interferences on assays from different manufacturers. Initial verification studies, followed up with ongoing quality control testing, can help a laboratory ensure the accuracy of haemolysis, icterus and lipaemia index results, as well as assist in managing any biases in index results from analysers from different manufacturers. Similarities, and variations, in the effect of haemolysis, icterus and lipaemia interference in assays from different manufacturers can often be predicted from the mechanism of interference. Nevertheless, interference testing is required to confirm expected similarities or to quantify differences. It is important that laboratories are familiar with a number of interference testing protocols and the particular strengths and weaknesses of each. A rigorous approach to all aspects of haemolysis, icterus and lipaemia interference testing allows the analytical progress in index measurement to be translated into improved patient care. PMID:27147624

  1. Antioxidants and the Comet assay.

    PubMed

    Cemeli, Eduardo; Baumgartner, Adolf; Anderson, Diana

    2009-01-01

    It is widely accepted that antioxidants, either endogenous or from the diet, play a key role in preserving health. They are able to quench radical species generated in situations of oxidative stress, either triggered by pathologies or xenobiotics, and they protect the integrity of DNA from genotoxicants. Nevertheless, there are still many compounds with unclear or unidentified prooxidant/antioxidant activities. This is of concern since there is an increase in the number of compounds synthesized or extracted from vegetables to which humans might be exposed. Despite the well-established protective effects of fruit and vegetables, the antioxidant(s) responsible have not all been clearly identified. There might also be alternative mechanisms contributing to the protective effects for which a comprehensive description is lacking. In the last two decades, the Comet assay has been extensively used for the investigation of the effects of antioxidants and many reports can be found in the literature. The Comet assay, a relatively fast, simple, and sensitive technique for the analysis of DNA damage in all cell types, has been applied for the screening of chemicals, biomonitoring and intervention studies. In the present review, several of the most well-known antioxidants are considered. These include: catalase, superoxide dismutase, glutathione peroxidase, selenium, iron chelators, melatonin, melanin, vitamins (A, B, C and E), carotenes, flavonoids, isoflavones, tea polyphenols, wine polyphenols and synthetic antioxidants. Investigations showing beneficial as well as non-beneficial properties of the antioxidants selected, either at the in vitro, ex vivo or in vivo level are discussed.

  2. Assay of potentially contaminated propellant

    SciTech Connect

    Koster, J.E.; Williams, H.E. III; Scott, W.S.

    1995-02-01

    One of the decontamination and decommissioning projects within DOD is demilitarization of an aging stockpile of munitions. A large portion of the stockpile contains depleted uranium (DU) as an armor piercing core and so these munitions must be assayed for the presence of uranium in other components. The assay method must be fast and preferably easy to implement. Presence of DU is indicated by its alpha decay. The alpha particles in turn produce ions in the ambient air. If a significant fraction of these ions can escape the quantity of propellant, the ions can be detected instead of the alpha particles. As a test of the feasibility of detecting alpha emissions from DU somewhere within a cartridge of propellant, the transmission of ions through layers of real propellant was measured. The propellant is in the form of graphite-coated cylindrical pellets. A 105nun cartridge was modified for use as a pellet chamber. A check source served as an ion source. The ion detector consisted of a grid held at 300V coupled to an ammeter. Results confirm that this is a promising technique for testing the propellant for the presence of DU quickly yet with sensitivity.

  3. Superconducting magnets

    SciTech Connect

    Not Available

    1994-08-01

    This report discusses the following topics on superconducting magnets: D19B and -C: The next steps for a record-setting magnet; D20: The push beyond 10 T: Beyond D20: Speculations on the 16-T regime; other advanced magnets for accelerators; spinoff applications; APC materials development; cable and cabling-machine development; and high-{Tc} superconductor at low temperature.

  4. Activation of Schwann cells in vitro by magnetic nanocomposites via applied magnetic field

    PubMed Central

    Liu, Zhongyang; Huang, Liangliang; Liu, Liang; Luo, Beier; Liang, Miaomiao; Sun, Zhen; Zhu, Shu; Quan, Xin; Yang, Yafeng; Ma, Teng; Huang, Jinghui; Luo, Zhuojing

    2015-01-01

    Schwann cells (SCs) are attractive seed cells in neural tissue engineering, but their application is limited by attenuated biological activities and impaired functions with aging. Therefore, it is important to explore an approach to enhance the viability and biological properties of SCs. In the present study, a magnetic composite made of magnetically responsive magnetic nanoparticles (MNPs) and a biodegradable chitosan–glycerophosphate polymer were prepared and characterized. It was further explored whether such magnetic nanocomposites via applied magnetic fields would regulate SC biological activities. The magnetization of the magnetic nanocomposite was measured by a vibrating sample magnetometer. The compositional characterization of the magnetic nanocomposite was examined by Fourier-transform infrared and X-ray diffraction. The tolerance of SCs to the magnetic fields was tested by flow-cytometry assay. The proliferation of cells was examined by a 5-ethynyl-2-deoxyuridine-labeling assay, a PrestoBlue assay, and a Live/Dead assay. Messenger ribonucleic acid of BDNF, GDNF, NT-3, and VEGF in SCs was assayed by quantitative real-time polymerase chain reaction. The amount of BDNF, GDNF, NT-3, and VEGF secreted from SCs was determined by enzyme-linked immunosorbent assay. It was found that magnetic nanocomposites containing 10% MNPs showed a cross-section diameter of 32.33±1.81 µm, porosity of 80.41%±0.72%, and magnetization of 5.691 emu/g at 8 kOe. The 10% MNP magnetic nanocomposites were able to support cell adhesion and spreading and further promote proliferation of SCs under magnetic field exposure. Interestingly, a magnetic field applied through the 10% MNP magnetic scaffold significantly increased the gene expression and protein secretion of BDNF, GDNF, NT-3, and VEGF. This work is the first stage in our understanding of how to precisely regulate the viability and biological properties of SCs in tissue-engineering grafts, which combined with additional

  5. Activation of Schwann cells in vitro by magnetic nanocomposites via applied magnetic field.

    PubMed

    Liu, Zhongyang; Huang, Liangliang; Liu, Liang; Luo, Beier; Liang, Miaomiao; Sun, Zhen; Zhu, Shu; Quan, Xin; Yang, Yafeng; Ma, Teng; Huang, Jinghui; Luo, Zhuojing

    2015-01-01

    Schwann cells (SCs) are attractive seed cells in neural tissue engineering, but their application is limited by attenuated biological activities and impaired functions with aging. Therefore, it is important to explore an approach to enhance the viability and biological properties of SCs. In the present study, a magnetic composite made of magnetically responsive magnetic nanoparticles (MNPs) and a biodegradable chitosan-glycerophosphate polymer were prepared and characterized. It was further explored whether such magnetic nanocomposites via applied magnetic fields would regulate SC biological activities. The magnetization of the magnetic nanocomposite was measured by a vibrating sample magnetometer. The compositional characterization of the magnetic nanocomposite was examined by Fourier-transform infrared and X-ray diffraction. The tolerance of SCs to the magnetic fields was tested by flow-cytometry assay. The proliferation of cells was examined by a 5-ethynyl-2-deoxyuridine-labeling assay, a PrestoBlue assay, and a Live/Dead assay. Messenger ribonucleic acid of BDNF, GDNF, NT-3, and VEGF in SCs was assayed by quantitative real-time polymerase chain reaction. The amount of BDNF, GDNF, NT-3, and VEGF secreted from SCs was determined by enzyme-linked immunosorbent assay. It was found that magnetic nanocomposites containing 10% MNPs showed a cross-section diameter of 32.33±1.81 µm, porosity of 80.41%±0.72%, and magnetization of 5.691 emu/g at 8 kOe. The 10% MNP magnetic nanocomposites were able to support cell adhesion and spreading and further promote proliferation of SCs under magnetic field exposure. Interestingly, a magnetic field applied through the 10% MNP magnetic scaffold significantly increased the gene expression and protein secretion of BDNF, GDNF, NT-3, and VEGF. This work is the first stage in our understanding of how to precisely regulate the viability and biological properties of SCs in tissue-engineering grafts, which combined with additional

  6. Data transformation methods for multiplexed assays

    DOEpatents

    Tammero, Lance F. Bentley; Dzenitis, John M; Hindson, Benjamin J

    2013-07-23

    Methods to improve the performance of an array assay are described. A correlation between fluorescence intensity-related parameters and negative control values of the assay is determined. The parameters are then adjusted as a function of the correlation. As a result, sensitivity of the assay is improved without changes in its specificity.

  7. Magnetizing of permanent magnets using HTS bulk magnets

    NASA Astrophysics Data System (ADS)

    Oka, Tetsuo; Muraya, Tomoki; Kawasaki, Nobutaka; Fukui, Satoshi; Ogawa, Jun; Sato, Takao; Terasawa, Toshihisa

    2012-01-01

    A demagnetized Nd-Fe-B permanent magnet was scanned just above the magnetic pole which contains the HTS bulk magnet generating a magnetic field of 3.27 T. The magnet sample was subsequently found to be fully magnetized in the open space of the static magnetic fields. We examined the magnetic field distributions when the magnetic poles were scanned twice to activate the magnet plate inversely with various overlap distances between the tracks of the bulk magnet. The magnetic field of the "rewritten" magnet reached the values of the magnetically saturated region of the material, showing steep gradients at the border of each magnetic pole. As a replacement for conventional pulse field magnetizing methods, this technique is proposed to expand the degree of freedom in the design of electromagnetic devices, and is proposed as a novel practical method for magnetizing rare-earth magnets, which have excellent magnetic performance and require intense fields of more than 3 T to be activated.

  8. Comparison of Established and Emerging Biodosimetry Assays

    PubMed Central

    Rothkamm, K.; Beinke, C.; Romm, H.; Badie, C.; Balagurunathan, Y.; Barnard, S.; Bernard, N.; Boulay-Greene, H.; Brengues, M.; De Amicis, A.; De Sanctis, S.; Greither, R.; Herodin, F.; Jones, A.; Kabacik, S.; Knie, T.; Kulka, U.; Lista, F.; Martigne, P.; Missel, A.; Moquet, J.; Oestreicher, U.; Peinnequin, A.; Poyot, T.; Roessler, U.; Scherthan, H.; Terbrueggen, B.; Thierens, H.; Valente, M.; Vral, A.; Zenhausern, F.; Meineke, V.; Braselmann, H.; Abend, M.

    2014-01-01

    Rapid biodosimetry tools are required to assist with triage in the case of a large-scale radiation incident. Here, we aimed to determine the dose-assessment accuracy of the well-established dicentric chromosome assay (DCA) and cytokinesis-block micronucleus assay (CBMN) in comparison to the emerging γ-H2AX foci and gene expression assays for triage mode biodosimetry and radiation injury assessment. Coded blood samples exposed to 10 X-ray doses (240 kVp, 1 Gy/min) of up to 6.4 Gy were sent to participants for dose estimation. Report times were documented for each laboratory and assay. The mean absolute difference (MAD) of estimated doses relative to the true doses was calculated. We also merged doses into binary dose categories of clinical relevance and examined accuracy, sensitivity and specificity of the assays. Dose estimates were reported by the first laboratories within 0.3–0.4 days of receipt of samples for the γ-H2AX and gene expression assays compared to 2.4 and 4 days for the DCA and CBMN assays, respectively. Irrespective of the assay we found a 2.5–4-fold variation of interlaboratory accuracy per assay and lowest MAD values for the DCA assay (0.16 Gy) followed by CBMN (0.34 Gy), gene expression (0.34 Gy) and γ-H2AX (0.45 Gy) foci assay. Binary categories of dose estimates could be discriminated with equal efficiency for all assays, but at doses ≥1.5 Gy a 10% decrease in efficiency was observed for the foci assay, which was still comparable to the CBMN assay. In conclusion, the DCA has been confirmed as the gold standard biodosimetry method, but in situations where speed and throughput are more important than ultimate accuracy, the emerging rapid molecular assays have the potential to become useful triage tools. PMID:23862692

  9. Human Arterial Ring Angiogenesis Assay.

    PubMed

    Seano, Giorgio; Primo, Luca

    2016-01-01

    In this chapter we describe a model of human angiogenesis where artery explants from umbilical cords are embedded in gel matrices and subsequently produce capillary-like structures. The human arterial ring (hAR) assay is an innovative system that enables three-dimensional (3D) and live studies of human angiogenesis. This ex vivo model has the advantage of recapitulating several steps of angiogenesis, including endothelial sprouting, migration, and differentiation into capillaries. Furthermore, it can be exploited for (1) identification of new genes regulating sprouting angiogenesis, (2) screening for pro- or anti-angiogenic drugs, (3) identification of biomarkers to monitor the efficacy of anti-angiogenic regimens, and (4) dynamic analysis of tumor microenvironmental effects on vessel formation. PMID:27172955

  10. Predictive Assay For Cancer Targets

    SciTech Connect

    Suess, A; Nguyen, C; Sorensen, K; Montgomery, J; Souza, B; Kulp, K; Dugan, L; Christian, A

    2005-09-19

    Early detection of cancer is a key element in successful treatment of the disease. Understanding the particular type of cancer involved, its origins and probable course, is also important. PhIP (2-amino-1-methyl-6 phenylimidazo [4,5-b]pyridine), a heterocyclic amine produced during the cooking of meat at elevated temperatures, has been shown to induce mammary cancer in female, Sprague-Dawley rats. Tumors induced by PhIP have been shown to contain discreet cytogenetic signature patterns of gains and losses using comparative genomic hybridization (CGH). To determine if a protein signature exists for these tumors, we are analyzing expression levels of the protein products of the above-mentioned tumors in combination with a new bulk protein subtractive assay. This assay produces a panel of antibodies against proteins that are either on or off in the tumor. Hybridization of the antibody panel onto a 2-D gel of tumor or control protein will allow for identification of a distinct protein signature in the tumor. Analysis of several gene databases has identified a number of rat homologs of human cancer genes located in these regions of gain and loss. These genes include the oncogenes c-MYK, ERBB2/NEU, THRA and tumor suppressor genes EGR1 and HDAC3. The listed genes have been shown to be estrogen-responsive, suggesting a possible link between delivery of bio-activated PhIP to the cell nucleus via estrogen receptors and gene-specific PhIP-induced DNA damage, leading to cell transformation. All three tumors showed similar silver staining patterns compared to each other, while they all were different than the control tissue. Subsequent screening of these genes against those from tumors know to be caused by other agents may produce a protein signature unique to PhIP, which can be used as a diagnostic to augment optical and radiation-based detection schemes.

  11. Development of forensic assay signatures for ebolaviruses.

    PubMed

    Song, Jian; Doggett, Norman; Wren, Melinda; Burr, Tom; Fenimore, P W; Hatcher, Eneida L; Bruno, William J; Li, Po-E; Stubben, Chris; Wolinsky, Murray

    2015-03-01

    Ebolaviruses are a diverse group of RNA viruses comprising five different species, four of which cause fatal hemorrhagic fever in humans. Because of their high infectivity and lethality, ebolaviruses are considered major biothreat agents. Although detection assays exist, no forensic assays are currently available. Here, we report the development of forensic assays that differentiate ebolaviruses. We performed phylogenetic analyses and identified canonical SNPs for all species, major clades and isolates. TaqMan-MGB allelic discrimination assays based on these SNPs were designed, screened against synthetic RNA templates, and validated against ebolavirus genomic RNAs. A total of 45 assays were validated to provide 100% coverage of the species and variants with additional resolution at the isolate level. These assays enabled accurate forensic analysis on 4 "unknown" ebolaviruses. Unknowns were correctly classified to species and variant. A goal of providing resolution below the isolate level was not successful. These high-resolution forensic assays allow rapid and accurate genotyping of ebolaviruses for forensic investigations.

  12. Magnetic investigations

    SciTech Connect

    Bath, G.D.; Jahren, C.E.; Rosenbaum, J.G.; Baldwin, M.J.

    1983-12-31

    Air and ground magnetic anomalies in the Climax stock area of the NTS help define the gross configuration of the stock and detailed configuration of magnetized rocks at the Boundary and Tippinip faults that border the stock. Magnetizations of geologic units were evaluated by measurements of magnetic properties of drill core, minimum estimates of magnetizations from ground magnetic anomalies for near surface rocks, and comparisons of measured anomalies with anomalies computed by a three-dimensional forward program. Alluvial deposits and most sedimentary rocks are nonmagnetic, but drill core measurements reveal large and irregular changes in magnetization for some quartzites and marbles. The magnetizations of quartz monzonite and granodiorite near the stock surface are weak, about 0.15 A/m, and increase at a rate of 0.00196 A/m/m to 1.55 A/m, at depths greater than 700 m (2300 ft). The volcanic rocks of the area are weakly magnetized. Aeromagnetic anomalies 850 m (2800 ft) above the stock are explained by a model consisting of five vertical prisms. Prisms 1, 2, and 3 represent the near surface outline of the stock, prism 4 is one of the models developed by Whitehill (1973), and prism 5 is modified from the model developed by Allingham and Zietz (1962). Most of the anomaly comes from unsampled and strongly-magnetized deep sources that could be either granite or metamorphosed sedimentary rocks. 48 refs., 23 figs., 3 tabs.

  13. Simple and sensitive microbial pathogen detection using a label-free DNA amplification assay.

    PubMed

    Sun, Yuhuan; Zhao, Chuanqi; Yan, Zhengqing; Ren, Jinsong; Qu, Xiaogang

    2016-06-14

    By the combination of quaternized magnetic nanoparticles and a label-free exonuclease III-assisted DNA amplification assay, we report a simple and facile strategy for the convenient and highly sensitive detection of microbial pathogens, with a detection limit of down to 50 cells mL(-1). PMID:27210898

  14. Immunomagnetic Reduction Assay on Des-Gamma-Carboxy Prothrombin for Screening of Hepatocellular Carcinoma.

    PubMed

    Chieh, Jen-Jie; Huang, K W; Chuang, C P; Wei, W C; Dong, J J; Lee, Y Y

    2016-08-01

    The accredited biomarker alpha-fetoprotein (AFP) offers limited sensitivity and specificity in the early detection of hepatocellular carcinoma (HCC). To improve the screening performance, des-gamma-carboxy prothrombin (DCP) has been identified as another promising biomarker of HCC, combined with AFP biomarkers. The results of the commercial optical enzyme-linked immunosorbent assay (ELISA) kit easily have the interference problem due to the optical methodology. The immunomagnetic reduction (IMR) assay based on the magnetic measurement was utilized to assay DCP biomarkers without the excellent antiinterference performances. A DCP magnetic reagent, composed of iron-oxide (Fe3O4 ) magnetic nanoparticles coated with anti-DCP antibodies solved in phosphoryl-buffer solution, was synthesized and characterized. In the test of standard DCP antigens, superior antiinterference and sensitivity than optical ELISA were proved. In the animal test, the results indicate good agreement between the IMR assay findings and the tumor sizes of HCC rats at all time points after the HCC implantation. The feasibility of the developed DCP magnetic reagent with the IMR for the detection of DCP is verified, and demonstrates the high potential for future clinical applications.

  15. Expert system for transuranic waste assay

    SciTech Connect

    Zoolalian, M.L.; Gibbs, A.; Kuhns, J.D.

    1989-01-01

    Transuranic wastes are generated at the Savannah River Site (SRS) as a result of routine production of nuclear materials. These wastes contain Pu-238 and Pu-239 and are placed into lined 55-gallon waste drums. The drums are placed on monitored storage pads pending shipment to the Waste Isolation Pilot Plant in New Mexico. A passive-active neutron (PAN) assay system is used to determine the mass of the radioactive material within the waste drums. Assay results are used to classify the wastes as either low-level or transuranic (TRU). During assays, the PAN assay system communicates with an IBM-AT computer. A Fortran computer program, called NEUT, controls and performs all data analyses. Unassisted, the NEUT program cannot adequately interpret assay results. To eliminate this limitation, an expert system shell was used to write a new algorithm, called the Transuranic Expert System (TRUX), to drive the NEUT program and add decision making capabilities for analysis of the assay results. The TRUX knowledge base was formulated by consulting with human experts in the field of neutron assay, by direct experimentation on the PAN assay system, and by observing operations on a daily basis. TRUX, with its improved ability to interpret assay results, has eliminated the need for close supervision by a human expert, allowing skilled technicians to operate the PAN assay system. 4 refs., 1 fig., 4 tabs.

  16. Planetary Magnetism

    NASA Technical Reports Server (NTRS)

    Connerney, J. E. P.

    2007-01-01

    The chapter on Planetary Magnetism by Connerney describes the magnetic fields of the planets, from Mercury to Neptune, including the large satellites (Moon, Ganymede) that have or once had active dynamos. The chapter describes the spacecraft missions and observations that, along with select remote observations, form the basis of our knowledge of planetary magnetic fields. Connerney describes the methods of analysis used to characterize planetary magnetic fields, and the models used to represent the main field (due to dynamo action in the planet's interior) and/or remnant magnetic fields locked in the planet's crust, where appropriate. These observations provide valuable insights into dynamo generation of magnetic fields, the structure and composition of planetary interiors, and the evolution of planets.

  17. Immunoliposome-based immunomagnetic concentration and separation assay for rapid detection of Cronobacter sakazakii.

    PubMed

    Shukla, Shruti; Lee, Gibaek; Song, Xinjie; Park, Sunhyun; Kim, Myunghee

    2016-03-15

    This study aimed to develop an immunoliposome-based immunomagnetic concentration and separation assay for the rapid detection of Cronobacter sakazakii (C. sakazakii), an acute opportunistic foodborne pathogenic bacterium, in both pure culture and infant formula. To develop the assay, magnetic nanoparticles (diameter 30 nm) were coated with immunoglobulin G (IgG), specifically anti-C. sakazakii IgG, and applied for the sensitive and efficient detection of C. sakazakii using immunoliposomes. The binding efficiency of anti-C. sakazakii IgG to the magnetic nanoparticles was 86.23 ± 0.59%. The assay developed in this study detected as few as 3.3 × 10(3) CFUmL(-1) of C. sakazakii in pure culture within 2h 30 min; in comparison, an indirect non-competitive enzyme-linked immunosorbent assay was able to detect 6.2 × 10(5) CFUmL(-1) of C. sakazakii in pure culture after 17 h. The developed assay did not show any cross-reactivity with other Cronobacter spp. or pathogens belonging to other genera. In addition, the method was able to detect 10(3) CFUmL(-1) of C. sakazakii in infant formula without any pre-incubation. These results confirm that the immunoliposome-based immunomagnetic concentration and separation assay may facilitate highly sensitive, efficient, and rapid detection of C. sakazakii. PMID:26547009

  18. Planetary Magnetism

    SciTech Connect

    Russell, C.T.

    1980-02-01

    Planetary spacecraft have now probed the magnetic fields of all the terrestrial planets, the moon, Jupiter, and Saturn. These measurements reveal that dynamos are active in at least four of the planets, Mercury, the earth, Jupiter, and Saturn but that Venus and Mars appear to have at most only very weak planetary magnetic fields. The moon may have once possessed an internal dynamo, for the surface rocks are magnetized. The large satellites of the outer solar system are candidates for dynamo action in addition to the large planets themselves. Of these satellites the one most likely to generate its own internal magnetic field is Io.

  19. Magnetic shielding

    DOEpatents

    Kerns, J.A.; Stone, R.R.; Fabyan, J.

    1987-10-06

    A magnetically-conductive filler material bridges the gap between a multi-part magnetic shield structure which substantially encloses a predetermined volume so as to minimize the ingress or egress of magnetic fields with respect to that volume. The filler material includes a heavy concentration of single-magnetic-domain-sized particles of a magnetically conductive material (e.g. soft iron, carbon steel or the like) dispersed throughout a carrier material which is generally a non-magnetic material that is at least sometimes in a plastic or liquid state. The maximum cross-sectional particle dimension is substantially less than the nominal dimension of the gap to be filled. An epoxy base material (i.e. without any hardening additive) low volatility vacuum greases or the like may be used for the carrier material. The structure is preferably exposed to the expected ambient magnetic field while the carrier is in a plastic or liquid state so as to facilitate alignment of the single-magnetic-domain-sized particles with the expected magnetic field lines. 3 figs.

  20. Magnetic shielding

    DOEpatents

    Kerns, John A.; Stone, Roger R.; Fabyan, Joseph

    1987-01-01

    A magnetically-conductive filler material bridges the gap between a multi-part magnetic shield structure which substantially encloses a predetermined volume so as to minimize the ingress or egress of magnetic fields with respect to that volume. The filler material includes a heavy concentration of single-magnetic-domain-sized particles of a magnetically conductive material (e.g. soft iron, carbon steel or the like) dispersed throughout a carrier material which is generally a non-magnetic material that is at least sometimes in a plastic or liquid state. The maximum cross-sectional particle dimension is substantially less than the nominal dimension of the gap to be filled. An epoxy base material (i.e. without any hardening additive) low volatility vacuum greases or the like may be used for the carrier material. The structure is preferably exposed to the expected ambient magnetic field while the carrier is in a plastic or liquid state so as to facilitate alignment of the single-magnetic-domain-sized particles with the expected magnetic field lines.

  1. Planetary magnetism

    NASA Technical Reports Server (NTRS)

    Russell, C. T.

    1980-01-01

    Planetary spacecraft have now probed the magnetic fields of all the terrestrial planets, the moon, Jupiter, and Saturn. These measurements reveal that dynamos are active in at least four of the planets, Mercury, the earth, Jupiter, and Saturn but that Venus and Mars appear to have at most only very weak planetary magnetic fields. The moon may have once possessed an internal dynamo, for the surface rocks are magnetized. The large satellites of the outer solar system are candidates for dynamo action in addition to the large planets themselves. Of these satellites the one most likely to generate its own internal magnetic field is Io.

  2. Automated amperometric plutonium assay system

    SciTech Connect

    Burt, M.C.

    1985-01-01

    The amperometric titration for plutonium assay has been used in the nuclear industry for over twenty years and has been in routine use at the Hanford Engineering Development Laboratory since 1976 for the analysis of plutonium oxide and mixed oxide fuel material for the Fast Flux Test Facility. It has proven itself to be an accurate and reliable method. The method may be used as a direct end point titration or an excess of titrant may be added and a back titration performed to aid in determination of the end point. Due to the slowness of the PuVI-FeII reaction it is difficult to recognize when the end point is being approached and is very time consuming if the current is allowed to decay to the residual value after each titrant addition. For this reason the back titration in which the rapid FeII-CrVI reaction occurs is used by most laboratories. The back titration is performed by the addition of excess ferrous solution followed by two measured aliquots of standard dichromate with measurement of cell current after each addition.

  3. Magneto immunofluorescence assay for diagnosis of celiac disease.

    PubMed

    Kergaravat, Silvina V; Beltramino, Luis; Garnero, Nidia; Trotta, Liliana; Wagener, Marta; Fabiano, Silvia N; Pividori, Maria Isabel; Hernandez, Silvia R

    2013-10-10

    A magneto immunofluorescence assay for the detection of anti-transglutaminase antibodies (ATG2) in celiac disease was developed. The ATG2 were recognized by transglutaminase enzyme immobilized on the magnetic beads and then the immunological reaction was revealed by antibodies labeled with peroxidase. The fluorescent response of the enzymatic reaction with o-phenylenediamine and H2O2 as substrates was correlated with anti-transglutaminase titer, showing EC50 and LOD values of 1:11,600 and 1:74,500 of antibody titers, respectively. A total number of 29 sera samples from clinically confirmed cases of celiac disease and 19 negative control samples were tested by the novel magneto immunofluorescence assay. The data were submitted to the receiver-operating characteristic plot (ROC) analysis which indicated that 8.1 U was the most effective cut-off value to discriminate correctly between celiac and non-celiac patients. The immunofluorescence assay exhibited a sensitivity of 96.6%, a specificity of 89.5% and an efficiency 93.8% compared with the commercial optical ELISA kit. PMID:24070488

  4. Magneto immunofluorescence assay for diagnosis of celiac disease.

    PubMed

    Kergaravat, Silvina V; Beltramino, Luis; Garnero, Nidia; Trotta, Liliana; Wagener, Marta; Fabiano, Silvia N; Pividori, Maria Isabel; Hernandez, Silvia R

    2013-10-10

    A magneto immunofluorescence assay for the detection of anti-transglutaminase antibodies (ATG2) in celiac disease was developed. The ATG2 were recognized by transglutaminase enzyme immobilized on the magnetic beads and then the immunological reaction was revealed by antibodies labeled with peroxidase. The fluorescent response of the enzymatic reaction with o-phenylenediamine and H2O2 as substrates was correlated with anti-transglutaminase titer, showing EC50 and LOD values of 1:11,600 and 1:74,500 of antibody titers, respectively. A total number of 29 sera samples from clinically confirmed cases of celiac disease and 19 negative control samples were tested by the novel magneto immunofluorescence assay. The data were submitted to the receiver-operating characteristic plot (ROC) analysis which indicated that 8.1 U was the most effective cut-off value to discriminate correctly between celiac and non-celiac patients. The immunofluorescence assay exhibited a sensitivity of 96.6%, a specificity of 89.5% and an efficiency 93.8% compared with the commercial optical ELISA kit.

  5. Assay development status report for total cyanide

    SciTech Connect

    Simpson, B.C.; Jones, T.E.; Pool, K.H.

    1993-02-01

    A validated cyanide assay that is applicable to a variety of tank waste matrices is necessary to resolve certain waste tank safety issues and for purposes of overall waste characterization. The target for this effort is an assay with an applicable range of greater than 1,000 ppM (0.10 wt%) total cyanide and a confidence level greater than 80%. Figure 1 illustrates the operating regime of the proposed cyanide assay method. The Assay Development Status Report for Total Cyanide will summarize the past experience with cyanide analyses on-tank waste matrices and will rate the status of the analytical methods used to assay total cyanide (CN{sup {minus}} ion) in the tank waste matrices as acceptable or unacceptable. This paper will also briefly describe the current efforts for improving analytical resolution of the assays and the attempts at speciation.

  6. High-resolution α-amylase assay combined with high-performance liquid chromatography-solid-phase extraction-nuclear magnetic resonance spectroscopy for expedited identification of α-amylase inhibitors: proof of concept and α-amylase inhibitor in cinnamon.

    PubMed

    Okutan, Leyla; Kongstad, Kenneth T; Jäger, Anna K; Staerk, Dan

    2014-11-26

    Type 2 diabetes affects millions of people worldwide, and new improved drugs or functional foods containing selective α-amylase inhibitors are needed for improved management of blood glucose. In this article the development of a microplate-based high-resolution α-amylase inhibition assay with direct photometric measurement of α-amylase activity is described. The inhibition assay is based on porcine pancreatic α-amylase with 2-chloro-4-nitrophenyl-α-D-maltotriose as substrate, which this gives a stable, sensitive, and cheap inhibition assay as requested for high-resolution purposes. In combination with HPLC-HRMS-SPE-NMR, this provides an analytical platform that allows simultaneous chemical and biological profiling of α-amylase inhibitors in plant extracts. Proof-of-concept with an artificial mixture of six compounds-of which three are known α-amylase inhibitors-showed that the high-resolution α-amylase inhibition profiles allowed detection of sub-microgram amounts of the α-amylase inhibitors. Furthermore, the high-resolution α-amylase inhibition assay/HPLC-HRMS-SPE-NMR platform allowed identification of cinnamaldehyde as the α-amylase inhibitor in cinnamon (Cinnamomum verum Presl.).

  7. Matrix effects of TRU (transuranic) assays using the SWEPP PAN assay system

    SciTech Connect

    Smith, J.R.

    1990-08-01

    The Drum Assay System (DAS) at the Stored Waste Experimental Pilot Plant (SWEPP) is a second-generation active-passive neutron assay system. It has been used to assay over 5000 208-liter drums of transuranic waste from the Rocky Flats Plant (RFP). Data from these assays have been examined and compared with the assays performed at Rocky Flats, mainly utilize counting of {sup 239}Pu gamma rays. For the most part the passive assays are in very good agreement with the Rocky Flats assays. The active assays are strongly correlated with the results of the other two methods, but require matrix-dependent correction factors beyond those provided by the system itself. A set of matrix-dependent correction factors has been developed from the study of the assay results. 3 refs., 4 figs., 3 tabs.

  8. Sensitive field assays for water analysis

    SciTech Connect

    Douglas, W.L.

    1984-08-01

    The goal of the project is to develop a rapid, simple, and inexpensive dry-film assay device for detection of environmental contaminants using the compound geosmin as a model. Phase I activities centered upon the immunochemical reagents necessary for the assay, development of an enzyme-cycling system that makes possible detection of substances in the parts per billion (PPB) range or lower, and demonstration of how the Immuno-Replacement-Assay can be used to detect geosmin.

  9. Multiplex assays to diagnose celiac disease.

    PubMed

    Lochman, Ivo; Martis, Peter; Burlingame, Rufus W; Lochmanová, Alexandra

    2007-08-01

    Patients with celiac disease are sensitive to the gluten fractions of wheat. Symptoms include gastrointestinal problems and a failure to thrive in children, but may range from headaches to general malaise in adults. Thus, it is difficult to diagnose celiac disease by symptoms alone. The standard diagnostic criteria include the presence of the characteristic anti-gliadin or anti-tissue transglutaminase antibodies (anti-tTG) in serum, flattened mucosa on intestinal biopsy, and improved symptoms on a gluten-free diet. Because of the ease of use of the tTG enzyme-linked immunosorbent assay (ELISA) compared to endomysial by indirect immunofluorescence assay, there has been much more screening for celiac disease in recent years. This increased screening showed that celiac disease was more prevalent than previously believed. We compared a new multiplex assay that includes a novel form of deamidated gliadin and recombinant human tTG as the antigens to other assays using standard antigens. In addition, the new assay detects the presence of selective IgA deficiency, which shows a 10-fold increase in prevalence in patients with celiac disease compared to the general population. The combination of sensitivity and specificity of the new multiplex assay was equal or better than those for standard assays. Thus the performance, ease of use, and ability to measure three clinically important parameters in a single test make the new multiplex assay a viable alternative to standard assays in a clinical lab.

  10. 233U Assay A Neutron NDA System

    SciTech Connect

    Hensley, D.C.; Lucero, A.J.; Pierce, L.

    1998-11-17

    The assay of highly enriched {sup 233}U material presents some unique challenges. Techniques which apply to the assay of materials of Pu or enriched {sup 235}U do not convert easily over to the assay of {sup 233}U. A specialized neutron assay device is being fabricated to exploit the singles neutron signal, the weak correlated neutron signal, and an active correlated signal. These pieces of information when combined with {gamma} ray isotopics information should give a good overall determination of {sup 233}U material now stored in bldg. 3019 at the Oak Ridge National Laboratory.

  11. Magnetic Recording.

    ERIC Educational Resources Information Center

    Lowman, Charles E.

    A guide to the technology of magnetic recorders used in such fields as audio recording, broadcast and closed-circuit television, instrumentation recording, and computer data systems is presented. Included are discussions of applications, advantages, and limitations of magnetic recording, its basic principles and theory of operation, and its…

  12. Magnetic shielding

    DOEpatents

    Kerns, J.A.; Stone, R.R.; Fabyan, J.

    1985-02-12

    A magnetically-conductive filler material bridges the gap between a multi-part magnetic shield structure which substantially encloses a predetermined volume so as to minimize the ingress or egress of magnetic fields with respect to that volume. The filler material includes a heavy concentration of single-magnetic-domain-sized particles of a magnetically conductive material (e.g. soft iron, carbon steel or the like) dispersed throughout a carrier material which is generally a non-magnetic material that is at least sometimes in a plastic or liquid state. The maximum cross-sectional particle dimension is substantially less than the nominal dimension of the gap to be filled. An epoxy base material (i.e. without any hardening additive) low volatility vacuum greases or the like may be used for the carrier material. The structure is preferably exposed to the expected ambient field while the carrier is in a plastic or liquid state so as to facilitate alignment of the single-magnetic-domain-sized particles with the expected magnetic field lines.

  13. Magnet Healing?

    NASA Astrophysics Data System (ADS)

    Finegold, Leonard

    2000-03-01

    Many people are convinced that static magnets—applied to their skin—will heal ills, and many businesses sell such magnets. The biophysics of such healing was reviewed [1] together with the general biophysics of static fields. Birds and insects do use the earth’s magnetic field for navigation. While insect and frog egg development can clearly be influenced by high fields (7 T and 17 T respectively), there is no experimental evidence that small magnetic fields (of less than 0.5 T) might heal, and much evidence that they cannot heal. A puzzle to the physics community is: How to show laypersons that simple magnets (very probably) do not heal, however attractive that idea might be. [1] L. Finegold, The Physics of "Alternative Medicine": Magnet Therapy, The Scientific Review of Alternative Medicine 3:26-33 (1999).

  14. Further comparison of primary hit identification by different assay technologies and effects of assay measurement variability.

    PubMed

    Wu, Xiang; Sills, Matthew A; Zhang, Ji-Hu

    2005-09-01

    High-throughput screening (HTS) has grown rapidly in the past decade, with many advances in new assay formats, detection technologies, and laboratory automation. Recently, several studies have shown that the choice of assay technology used for the screening process is particularly important and can yield quite different primary screening outcomes. However, because the screening assays in these previous studies were performed in a single-point determination, it is not clear to what extent the difference observed in the screening results between different assay technologies is attributable to inherent assay variability and day-to-day measurement variation. To address this question, a nuclear receptor coactivator recruitment assay was carried out in 2 different assay formats, namely, AlphaScreen and time-resolved fluorescence resonance energy transfer, which probed the same biochemical binding events but with different detection technologies. For each assay format, 4 independent screening runs in a typical HTS setting were completed to evaluate the run-to-run screening variability. These multiple tests with 2 assay formats allow an unambiguous comparison between the discrepancies of different assay formats and the effects of the variability of assay and screening measurements on the screening outcomes. The results provide further support that the choice of assay format or technology is a critical factor in HTS assay development.

  15. 21 CFR 864.7525 - Heparin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification....

  16. 21 CFR 864.7525 - Heparin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification....

  17. 21 CFR 864.7525 - Heparin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification....

  18. 21 CFR 864.7525 - Heparin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification....

  19. 21 CFR 864.7525 - Heparin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification....

  20. Statistical inference for serial dilution assay data.

    PubMed

    Lee, M L; Whitmore, G A

    1999-12-01

    Serial dilution assays are widely employed for estimating substance concentrations and minimum inhibitory concentrations. The Poisson-Bernoulli model for such assays is appropriate for count data but not for continuous measurements that are encountered in applications involving substance concentrations. This paper presents practical inference methods based on a log-normal model and illustrates these methods using a case application involving bacterial toxins.

  1. A simplified plaque assay for varicella vaccine.

    PubMed

    Husson-van Vliet, J; Colinet, G; Yane, F; Lemoine, P

    1987-11-01

    A simple and accurate plaque assay is described for potency testing of attenuated varicella vaccine. Assays were performed on the African green monkey kidney continuous cell line CV-1, in multidish-plates, under a semi-solid carboxymethylcellulose overlay. The test is economical and yields accurate individual titre estimates, the reliability of which may be assessed by parallel titration of reference preparations.

  2. 21 CFR 866.3210 - Endotoxin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay....

  3. 21 CFR 866.3210 - Endotoxin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay....

  4. 21 CFR 866.3210 - Endotoxin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay....

  5. 21 CFR 866.3210 - Endotoxin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay....

  6. 21 CFR 866.3210 - Endotoxin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay....

  7. Development of an upconverting chelate assay

    NASA Astrophysics Data System (ADS)

    Xiao, Xudong; Haushalter, Jeanne P.; Kotz, Kenneth T.; Faris, Gregory W.

    2005-04-01

    We report progress on performing a cell-based assay for the detection of EGFR on cell surfaces by using upconverting chelates. An upconversion microscope has been developed for performing assays and testing optical response. A431 cells are labeled with europium DOTA and imaged using this upconverting microscope.

  8. Pulling on super paramagnetic beads with micro cantilevers: single molecule mechanical assay application.

    PubMed

    Muñoz, Romina; Aguilar Sandoval, Felipe; Wilson, Christian A M; Melo, Francisco

    2015-07-22

    This paper demonstrates that it is possible to trap and release a super paramagnetic micro bead by fixing three super paramagnetic micro beads in a triangular array at the sensitive end of a micro cantilever, and by simply switching on/off an external magnetic field. To provide evidence of this principle we trap a micro bead that is attached to the free end of single DNA molecule and that has been previously fixed at the other end to a glass surface, using the standard sample preparation protocol of magnetic tweezers assays. The switching process is reversible which preserves the integrity of the tethered molecule, and a local force applied over the tethered bead excludes the neighbouring beads from the magnetic trap. We have developed a quadrature phase interferometer which is able to perform under fluid environments to accurately measure small deflections, which permits the exploration of DNA elasticity. Our results agree with measurements from magnetic tweezer assays performed under similar conditions. Furthermore, compared to the magnetic tweezer methodology, the combination of the magnetic trap with a suitable measurement system for cantilever deflection, allows for the exploration of a wide range of forces using a local method that has an improved temporal resolution.

  9. Pulling on super paramagnetic beads with micro cantilevers: single molecule mechanical assay application

    NASA Astrophysics Data System (ADS)

    Muñoz, Romina; Aguilar Sandoval, Felipe; Wilson, Christian A. M.; Melo, Francisco

    2015-07-01

    This paper demonstrates that it is possible to trap and release a super paramagnetic micro bead by fixing three super paramagnetic micro beads in a triangular array at the sensitive end of a micro cantilever, and by simply switching on/off an external magnetic field. To provide evidence of this principle we trap a micro bead that is attached to the free end of single DNA molecule and that has been previously fixed at the other end to a glass surface, using the standard sample preparation protocol of magnetic tweezers assays. The switching process is reversible which preserves the integrity of the tethered molecule, and a local force applied over the tethered bead excludes the neighbouring beads from the magnetic trap. We have developed a quadrature phase interferometer which is able to perform under fluid environments to accurately measure small deflections, which permits the exploration of DNA elasticity. Our results agree with measurements from magnetic tweezer assays performed under similar conditions. Furthermore, compared to the magnetic tweezer methodology, the combination of the magnetic trap with a suitable measurement system for cantilever deflection, allows for the exploration of a wide range of forces using a local method that has an improved temporal resolution.

  10. Acellular comet assay: a tool for assessing variables influencing the alkaline comet assay.

    PubMed

    Kennedy, Erin K; McNamee, James P; Prud'homme Lalonde, Louise; Jones, Trevor; Wilkinson, Diana

    2012-01-01

    In this study, an acellular modification to the alkaline comet assay to further evaluate key variables within the assay that may influence the outcome of genotoxicity studies is described. This acellular comet assay can detect differences of 0.2 Gy of (60)Co gamma-ray radiation between 0 and 1 Gy and differences of 1 Gy between 0 and 8 Gy; thus, this assay is applicable for a wide range of DNA damage levels. It is also shown that DNA damage from different radiation energies was not significantly different from (60)Co gamma-ray. This assay displayed a statistical increase in DNA damage due to uncontrolled exposure to natural light; however, the slope of the dose-response curve for light-exposed samples was similar to that for samples protected from light. A comparison of the alkaline comet assay with the acellular comet assay allowed for the intrinsic repair capacity of the alkaline comet assay to be quantified.

  11. A highly sensitive telomerase activity assay that eliminates false-negative results caused by PCR inhibitors.

    PubMed

    Yaku, Hidenobu; Murashima, Takashi; Miyoshi, Daisuke; Sugimoto, Naoki

    2013-01-01

    An assay for telomerase activity based on asymmetric polymerase chain reaction (A-PCR) on magnetic beads (MBs) and subsequent application of cycling probe technology (CPT) is described. In this assay, the telomerase reaction products are immobilized on MBs, which are then washed to remove PCR inhibitors that are commonly found in clinical samples. The guanine-rich sequences (5'-(TTAGGG)n-3') of the telomerase reaction products are then preferentially amplified by A-PCR, and the amplified products are subsequently detected via CPT, where a probe RNA with a fluorophore at the 5' end and a quencher at the 3' end is hydrolyzed by RNase H in the presence of the target DNA. The catalyst-mediated cleavage of the probe RNA enhances fluorescence from the 5' end of the probe. The assay allowed us to successfully detect HeLa cells selectively over normal human dermal fibroblast (NHDF) cells. Importantly, this selectivity produced identical results with regard to detection of HeLa cells in the absence and presence of excess NHDF cells; therefore, this assay can be used for practical clinical applications. The lower limit of detection for HeLa cells was 50 cells, which is lower than that achieved with a conventional telomeric repeat amplification protocol assay. Our assay also eliminated false-negative results caused by PCR inhibitors. Furthermore, we show that this assay is appropriate for screening among G-quadruplex ligands to find those that inhibit telomerase activity.

  12. A highly sensitive telomerase activity assay that eliminates false-negative results caused by PCR inhibitors.

    PubMed

    Yaku, Hidenobu; Murashima, Takashi; Miyoshi, Daisuke; Sugimoto, Naoki

    2013-01-01

    An assay for telomerase activity based on asymmetric polymerase chain reaction (A-PCR) on magnetic beads (MBs) and subsequent application of cycling probe technology (CPT) is described. In this assay, the telomerase reaction products are immobilized on MBs, which are then washed to remove PCR inhibitors that are commonly found in clinical samples. The guanine-rich sequences (5'-(TTAGGG)n-3') of the telomerase reaction products are then preferentially amplified by A-PCR, and the amplified products are subsequently detected via CPT, where a probe RNA with a fluorophore at the 5' end and a quencher at the 3' end is hydrolyzed by RNase H in the presence of the target DNA. The catalyst-mediated cleavage of the probe RNA enhances fluorescence from the 5' end of the probe. The assay allowed us to successfully detect HeLa cells selectively over normal human dermal fibroblast (NHDF) cells. Importantly, this selectivity produced identical results with regard to detection of HeLa cells in the absence and presence of excess NHDF cells; therefore, this assay can be used for practical clinical applications. The lower limit of detection for HeLa cells was 50 cells, which is lower than that achieved with a conventional telomeric repeat amplification protocol assay. Our assay also eliminated false-negative results caused by PCR inhibitors. Furthermore, we show that this assay is appropriate for screening among G-quadruplex ligands to find those that inhibit telomerase activity. PMID:24071983

  13. Dynamic, Quantitative Assays of Phagosomal Function

    PubMed Central

    Podinovskaia, Maria; VanderVen, Brian C.; Yates, Robin M.; Glennie, Sarah; Fullerton, Duncan; Mwandumba, Henry C.; Russell, David G.

    2013-01-01

    Much of the activity of the macrophage as an effector cell is performed within its phagocytic compartment. This ranges from the degradation of tissue debris as part of its homeostatic function, to the generation of the superoxide burst as part of its microbicidal response to infection. We have developed a range of real-time readouts of phagosomal function that enables these activities to be rigorously quantified. This chapter contains the description of several of these assays assessed by different methods of quantitation; including a Fluorescence Resonance Emission Transfer (FRET) assay for phagosome/lysosome fusion measured by spectrofluorometer, a fluorogenic assay for the superoxide burst measured by flow cytometry, and a fluorogenic assay for bulk proteolysis measure by confocal microscope. These assays illustrate both the range parameters that can be quantified as well as the flexibility of instrumentation that can be exploited for their quantitation. PMID:24510516

  14. Paper disk assay for glycosaminoglycan sulfotransferases

    SciTech Connect

    Sugahara, K.; Ishii, T.; Yamashina, I.

    1987-11-01

    A method is described for the assay of sulfotransferases, which transfer sulfate from 3'-phosphoadenosine-5'-phosphosulfate (PAPS) to glycosaminoglycan acceptors. Following the sulfation reactions, the (/sup 35/S)sulfate-labeled products are precipitated and then separated from a sulfate donor ((/sup 35/S)PAPS) and its degradation products by a paper disk method, and then the radioactivity remaining on the paper disk is subsequently determined by liquid scintillation counting. The rapidity and simplicity of the method are advantageous for multiple assays and have allowed us to establish assay conditions for serum sulfotransferases which introduce sulfate at position 6 of the internal N-acetylgalactosamine units of chondroitin, position 2 (amino group) of the glucosamine units of heparan sulfate and sugar units of keratan sulfate, respectively. The assay method will be applicable with modification to the assay of other glycosaminoglycan sulfotransferases and glycoprotein sulfotransferases.

  15. A Novel Detection Platform for Shrimp White Spot Syndrome Virus Using an ICP11-Dependent Immunomagnetic Reduction (IMR) Assay

    PubMed Central

    Liu, Bing-Hsien; Lin, Yu-Chen; Ho, Chia-Shin; Yang, Che-Chuan; Chang, Yun-Tsui; Chang, Jui-Feng; Li, Chun-Yuan; Cheng, Cheng-Shun; Huang, Jiun-Yan; Lee, Yen-Fu; Hsu, Ming-Hung; Lin, Feng-Chun; Wang, Hao-Ching; Lo, Chu-Fang; Yang, Shieh-Yueh; Wang, Han-Ching

    2015-01-01

    Shrimp white spot disease (WSD), which is caused by white spot syndrome virus (WSSV), is one of the world’s most serious shrimp diseases. Our objective in this study was to use an immunomagnetic reduction (IMR) assay to develop a highly sensitive, automatic WSSV detection platform targeted against ICP11 (the most highly expressed WSSV protein). After characterizing the magnetic reagents (Fe3O4 magnetic nanoparticles coated with anti ICP11), the detection limit for ICP11 protein using IMR was approximately 2 x 10−3 ng/ml, and the linear dynamic range of the assay was 0.1~1 x 106 ng/ml. In assays of ICP11 protein in pleopod protein lysates from healthy and WSSV-infected shrimp, IMR signals were successfully detected from shrimp with low WSSV genome copy numbers. We concluded that this IMR assay targeting ICP11 has potential for detecting the WSSV. PMID:26380977

  16. Hyaluronidase treatment of synovial fluid to improve assay precision for biomarker research using multiplex immunoassay platforms.

    PubMed

    Jayadev, Chethan; Rout, Raj; Price, Andrew; Hulley, Philippa; Mahoney, David

    2012-12-14

    Synovial fluid (SF) is a difficult biological matrix to analyse due to its complex non-Newtonian nature. This can result in poor assay repeatability and potentially inefficient use of precious samples. This study assessed the impact of SF treatment by hyaluronidase and/or dilution on intra-assay precision using the Luminex and Meso Scale Discovery (MSD) multiplex platforms. SF was obtained from patients with knee osteoarthritis at the time of joint replacement surgery. Aliquots derived from the same sample were left untreated (neat), 2-fold diluted, 4-fold diluted or treated with 2mg/ml testicular hyaluronidase (with 2-fold dilution). Preparation methods were compared in a polysterene-bead Luminex 10-plex (N=16), magnetic-bead Luminex singleplex (N=7) and MSD 4-plex (N=7). Each method was assessed for coefficient of variation (CV) of replicate measurements, number of bead events (for Luminex assays) and dilution-adjusted analyte concentration. Percentage recovery was calculated for dilutions and HAse treatment. Hyaluronidase treatment significantly increased the number of wells with satisfactory bead events/region (95%) compared to neat (48%, p<0.001) in the polystyrene-bead Luminex assay, but the magnetic-bead Luminex assay achieved ≥50 bead events irrespective of treatment method. Hyaluronidase treatment resulted in lower intra-assay CVs for detectable ligands (group average CV<10%) than neat, 2-fold and 4-fold dilution (CV~25% for all, p<0.05) in both polystyrene- and magnetic-bead Luminex assays. In addition, measured sample concentrations were higher and recovery was poor (elevated) after hyaluronidase treatment. In the MSD 4-plex, within-group comparison of the intra-assay CV or concentration was not conclusively influenced by SF preparation. However, only hyaluronidase treatment resulted in CV<25% for all samples for TNF-α. There was no effect on analyte concentrations or recovery. Hyaluronidase treatment can improve intra-assay precision and assay signal

  17. Magnetic Bearing

    NASA Technical Reports Server (NTRS)

    1996-01-01

    AVCON, Inc. produces advanced magnetic bearing systems for industrial use, offering a unique technological approach based on contract work done at Marshall Space Flight Center and Lewis Research Center. Designed for the turbopump of the Space Shuttle main engine, they are now used in applications such as electric power generation, petroleum refining, machine tool operation and natural gas pipelines. Magnetic bearings support moving machinery without physical contact; AVCON's homopolar approach is a hybrid of permanent and electromagnets which are one-third the weight, smaller and more power- efficient than previous magnetic bearings.

  18. Lunar magnetism

    NASA Technical Reports Server (NTRS)

    Hood, L. L.; Sonett, C. P.; Srnka, L. J.

    1984-01-01

    Aspects of lunar paleomagnetic and electromagnetic sounding results which appear inconsistent with the hypothesis that an ancient core dynamo was the dominant source of the observed crustal magnetism are discussed. Evidence is summarized involving a correlation between observed magnetic anomalies and ejecta blankets from impact events which indicates the possible importance of local mechanisms involving meteoroid impact processes in generating strong magnetic fields at the lunar surface. A reply is given to the latter argument which also presents recent evidence of a lunar iron core.

  19. Performance Evaluation of the Serum Thyroglobulin Assays With Immunochemiluminometric Assay and Immunoradiometric Assay for Differentiated Thyroid Cancer

    PubMed Central

    Cho, Yoon Young; Chun, Sejong; Lee, Soo-Youn; Chung, Jae Hoon

    2016-01-01

    Background Measurement of postoperative serum thyroglobulin (Tg) is important for detecting persistent or recurrent differentiated thyroid cancer. We evaluated the analytic performance of the DxI 800 assay (Beckman Coulter, USA) for serum Tg and anti-thyroglobulin antibodies (TgAbs) in comparison with that of the GAMMA-10 assay (Shinjin Medics Inc., Korea) for serum Tg and RIA-MAT 280 assay (Stratec, Germany) for TgAb. Methods We prospectively collected blood samples from 99 patients thyroidectomized for thyroid cancer. The functional sensitivity was investigated in standards and human serum. Precision and linearity were evaluated according to the guidelines of the Clinical and Laboratory Standards Institute. The correlation between the two assays was assessed in samples with different Tg ranges. Results The functional sensitivity of the DxI 800 assay for serum Tg was between 0.0313 and 0.0625 ng/mL. The total CV was 3.9–5.6% for serum Tg and 5.3–6.9% for serum TgAb. The coefficient of determination (R2) was 1.0 and 0.99 for serum Tg and TgAb, respectively. The cut-offs for serum TgAb were 4.0 IU/mL (DxI 800) and 60.0 IU/mL (RIA-MAT 280), and the overall agreement was 68.7%. The correlation between the two assays was excellent; the correlation coefficient was 0.99 and 0.88 for serum Tg and TgAb, respectively. Conclusions The DxI 800 is a sensitive assay for serum Tg and TgAb, and the results correlated well with those from the immunoradiometric assays (IRMA). This assay has several advantages over the IRMA and could be considered an alternative test for Tg measurement. PMID:27374705

  20. Imaging Intelligence with Proton Magnetic Resonance Spectroscopy

    ERIC Educational Resources Information Center

    Jung, Rex E.; Gasparovic, Charles; Chavez, Robert S.; Caprihan, Arvind; Barrow, Ranee; Yeo, Ronald A.

    2009-01-01

    Proton magnetic resonance spectroscopy ([to the first power]H-MRS) is a technique for the assay of brain neurochemistry "in vivo." N-acetylaspartate (NAA), the most prominent metabolite visible within the [to the first power]H-MRS spectrum, is found primarily within neurons. The current study was designed to further elucidate NAA-cognition…

  1. Ultrarapid Detection of Pathogenic Bacteria Using a 3D Immunomagnetic Flow Assay

    PubMed Central

    Lee, Wonjae; Kwon, Donghoon; Chung, Boram; Jung, Gyoo Yeol; Au, Anthony; Folch, Albert; Jeon, Sangmin

    2015-01-01

    We developed a novel 3D immunomagnetic flow assay for the rapid detection of pathogenic bacteria in a large-volume food sample. Antibody-functionalized magnetic nanoparticle clusters (AbMNCs) were magnetically immobilized on the surfaces of a 3D-printed cylindrical microchannel. The injection of a Salmonella-spiked sample solution into the microchannel produced instant binding between the AbMNCs and the Salmonella bacteria due to their efficient collisions. Nearly perfect capture of the AbMNCs and AbMNCs-Salmonella complexes was achieved under a high flow rate by stacking permanent magnets with spacers inside the cylindrical separator to maximize the magnetic force. The concentration of the bacteria in solution was determined using ATP luminescence measurements. The detection limit was better than 10 cfu/mL, and the overall assay time, including the binding, rinsing, and detection steps for a 10 mL sample took less than 3 min. To our knowledge, the 3D immunomagnetic flow assay described here provides the fastest high-sensitivity, high-capacity method for the detection of pathogenic bacteria. PMID:24856003

  2. Ultrarapid detection of pathogenic bacteria using a 3D immunomagnetic flow assay.

    PubMed

    Lee, Wonjae; Kwon, Donghoon; Chung, Boram; Jung, Gyoo Yeol; Au, Anthony; Folch, Albert; Jeon, Sangmin

    2014-07-01

    We developed a novel 3D immunomagnetic flow assay for the rapid detection of pathogenic bacteria in a large-volume food sample. Antibody-functionalized magnetic nanoparticle clusters (AbMNCs) were magnetically immobilized on the surfaces of a 3D-printed cylindrical microchannel. The injection of a Salmonella-spiked sample solution into the microchannel produced instant binding between the AbMNCs and the Salmonella bacteria due to their efficient collisions. Nearly perfect capture of the AbMNCs and AbMNCs-Salmonella complexes was achieved under a high flow rate by stacking permanent magnets with spacers inside the cylindrical separator to maximize the magnetic force. The concentration of the bacteria in solution was determined using ATP luminescence measurements. The detection limit was better than 10 cfu/mL, and the overall assay time, including the binding, rinsing, and detection steps for a 10 mL sample took less than 3 min. To our knowledge, the 3D immunomagnetic flow assay described here provides the fastest high-sensitivity, high-capacity method for the detection of pathogenic bacteria.

  3. Magnetic Reconnection

    NASA Video Gallery

    This science visualization shows a magnetospheric substorm, during which, magnetic reconnection causes energy to be rapidly released along the field lines in the magnetotail, that part of the magne...

  4. Magnetic Bacteria.

    ERIC Educational Resources Information Center

    Nelson, Jane Bray; Nelson, Jim

    1992-01-01

    Describes the history of Richard Blakemore's discovery of magnetotaxic organisms. Discusses possible reasons why the magnetic response in bacteria developed. Proposes research experiments integrating biology and physics in which students investigate problems using cultures of magnetotaxic organisms. (MDH)

  5. Magnetic monopoles

    SciTech Connect

    Fryberger, D.

    1984-12-01

    In this talk on magnetic monopoles, first the author briefly reviews some historical background; then, the author describes what several different types of monopoles might look like; and finally the author discusses the experimental situation. 81 references.

  6. CRYOGENIC MAGNETS

    DOEpatents

    Post, R.F.; Taylor, C.E.

    1963-05-21

    A cryogenic magnet coil is described for generating magnetic fields of the order of 100,000 gauss with a minimum expenditure of energy lost in resistive heating of the coil inductors and energy lost irreversibly in running the coil refrigeration plant. The cryogenic coil comprises a coil conductor for generating a magnetic field upon energization with electrical current, and refrigeration means disposed in heat conductive relation to the coil conductor for cooling to a low temperature. A substantial reduction in the power requirements for generating these magnetic fields is attained by scaling the field generating coil to large size and particular dimensions for a particular conductor, and operating the coil at a particular optimum temperature commensurate with minimum overall power requirements. (AEC)

  7. Detection of magnetic resonance signals using a magnetoresistive sensor

    SciTech Connect

    Budker, Dmitry; Pines, Alexander; Xu, Shoujun; Hilty, Christian; Ledbetter, Micah P; Bouchard, Louis S

    2013-10-01

    A method and apparatus are described wherein a micro sample of a fluidic material may be assayed without sample contamination using NMR techniques, in combination with magnetoresistive sensors. The fluidic material to be assayed is first subject to pre-polarization, in one embodiment, by passage through a magnetic field. The magnetization of the fluidic material is then subject to an encoding process, in one embodiment an rf-induced inversion by passage through an adiabatic fast-passage module. Thereafter, the changes in magnetization are detected by a pair of solid-state magnetoresistive sensors arranged in gradiometer mode. Miniaturization is afforded by the close spacing of the various modules.

  8. DNA Methyltransferase Activity Assays: Advances and Challenges

    PubMed Central

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao, Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modification, clinical diagnosis and therapeutics. Conventional techniques such as isotope labelling are effective, but they often require laborious sample preparation, isotope labelling, sophisticated equipment and large amounts of DNA, rendering them unsuitable for uses at point-of-care. Simple, portable, highly sensitive and low-cost assays are urgently needed for DNA MTase activity screening. In most recent technological advances, many alternative DNA MTase activity assays such as fluorescent, electrochemical, colorimetric and chemiluminescent assays have been proposed. In addition, many of them are coupled with nanomaterials and/or enzymes to significantly enhance their sensitivity. Herein we review the progress in the development of DNA MTase activity assays with an emphasis on assay mechanism and performance with some discussion on challenges and perspectives. It is hoped that this article will provide a broad coverage of DNA MTase activity assays and their latest developments and open new perspectives toward the development of DNA MTase activity assays with much improved performance for uses in molecular biology and clinical practice. PMID:26909112

  9. Nano-immunosafety: issues in assay validation

    NASA Astrophysics Data System (ADS)

    Boraschi, Diana; Oostingh, Gertie J.; Casals, Eudald; Italiani, Paola; Nelissen, Inge; Puntes, Victor F.; Duschl, Albert

    2011-07-01

    Assessing the safety of engineered nanomaterials for human health must include a thorough evaluation of their effects on the immune system, which is responsible for defending the integrity of our body from damage and disease. An array of robust and representative assays should be set up and validated, which could be predictive of the effects of nanomaterials on immune responses. In a trans-European collaborative work, in vitro assays have been developed to this end. In vitro tests have been preferred for their suitability to standardisation and easier applicability. Adapting classical assays to testing the immunotoxicological effects of nanoparticulate materials has raised a series of issues that needed to be appropriately addressed in order to ensure reliability of results. Besides the exquisitely immunological problem of selecting representative endpoints predictive of the risk of developing disease, assay results turned out to be significantly biased by artefactual interference of the nanomaterials or contaminating agents with the assay protocol. Having addressed such problems, a series of robust and representative assays have been developed that describe the effects of engineered nanoparticles on professional and non-professional human defence cells. Two of such assays are described here, one based on primary human monocytes and the other employing human lung epithelial cells transfected with a reporter gene.

  10. DNA Methyltransferase Activity Assays: Advances and Challenges.

    PubMed

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao, Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modification, clinical diagnosis and therapeutics. Conventional techniques such as isotope labelling are effective, but they often require laborious sample preparation, isotope labelling, sophisticated equipment and large amounts of DNA, rendering them unsuitable for uses at point-of-care. Simple, portable, highly sensitive and low-cost assays are urgently needed for DNA MTase activity screening. In most recent technological advances, many alternative DNA MTase activity assays such as fluorescent, electrochemical, colorimetric and chemiluminescent assays have been proposed. In addition, many of them are coupled with nanomaterials and/or enzymes to significantly enhance their sensitivity. Herein we review the progress in the development of DNA MTase activity assays with an emphasis on assay mechanism and performance with some discussion on challenges and perspectives. It is hoped that this article will provide a broad coverage of DNA MTase activity assays and their latest developments and open new perspectives toward the development of DNA MTase activity assays with much improved performance for uses in molecular biology and clinical practice.

  11. Global haemostasis assays, from bench to bedside.

    PubMed

    van Geffen, Mark; van Heerde, Waander L

    2012-06-01

    Bleeding and thrombosis are the ultimate clinical outcomes of aberrations in the haemostatic process. Haemostasis prevents excessive blood loss due to the effort of various compartments like the vasculature, blood cells, coagulation and fibrinolysis. The complexity of all processes involved makes the diagnosis of aberrations difficult, cumbersome and expensive. A single assay to detect any factor disturbing this haemostatic balance with high sensitivity and specificity would be of great value, especially if the outcome of this assay correlates well with clinical outcome. Despite years of research, such an assay is not yet available; however, some interesting candidates are under development and combine the effects of various compartments. This review describes the development of global haemostasis assays and summarizes the current state of the art of these haemostasis assays covering thrombin and plasmin generation, turbidity and thromboelastography/thromboelastometry. Finally, we discuss the applicability of global assays in clinical practice and we provide a future perspective on the ongoing development of automation and miniaturisation as it is our belief that these developments will benefit the standardization of global haemostasis assays.

  12. The root causes of pharmacodynamic assay failure.

    PubMed

    Ferry-Galow, Katherine V; Makhlouf, Hala R; Wilsker, Deborah F; Lawrence, Scott M; Pfister, Thomas D; Marrero, Allison M; Bigelow, Kristina M; Yutzy, William H; Ji, Jiuping J; Butcher, Donna O; Gouker, Brad A; Kummar, Shivaani; Chen, Alice P; Kinders, Robert J; Parchment, Ralph E; Doroshow, James H

    2016-08-01

    Robust pharmacodynamic assay results are valuable for informing go/no-go decisions about continued development of new anti-cancer agents and for identifying combinations of targeted agents, but often pharmacodynamic results are too incomplete or variable to fulfill this role. Our experience suggests that variable reagent and specimen quality are two major contributors to this problem. Minimizing all potential sources of variability in procedures for specimen collection, processing, and assay measurements is essential for meaningful comparison of pharmacodynamic biomarkers across sample time points. This is especially true in the evaluation of pre- and post-dose tumor biopsies, which suffer from high levels of tumor insufficiency due to variations in biopsy collection techniques and significant specimen heterogeneity within and across patients. Developing methods to assess heterogeneous biopsies is necessary in order to evaluate a majority of tumor biopsies collected for pharmacodynamic biomarker studies. Improved collection devices and standardization of methods are being sought in order to improve the tumor content and quality of tumor biopsies. In terms of reagent variability, we have found that stringent initial reagent qualification and quality control of R&D-grade reagents is critical to minimize lot-to-lot variability and prevent assay failures, especially for clinical pharmacodynamic questions, which often demand assay performance that meets or exceeds clinical diagnostic assay standards. Rigorous reagent specifications and use of appropriate assay quality control methodologies help to ensure consistency between assay runs, laboratories and trials to provide much needed pharmacodynamic insights into the activity of investigational agents. PMID:27663480

  13. Superconducting magnet

    NASA Technical Reports Server (NTRS)

    1985-01-01

    Extensive computer based engineering design effort resulted in optimization of a superconducting magnet design with an average bulk current density of approximately 12KA/cm(2). Twisted, stranded 0.0045 inch diameter NbTi superconductor in a copper matrix was selected. Winding the coil from this bundle facilitated uniform winding of the small diameter wire. Test coils were wound using a first lot of the wire. The actual packing density was measured from these. Interwinding voltage break down tests on the test coils indicated the need for adjustment of the wire insulation on the lot of wire subsequently ordered for construction of the delivered superconducting magnet. Using the actual packing densities from the test coils, a final magnet design, with the required enhancement and field profile, was generated. All mechanical and thermal design parameters were then also fixed. The superconducting magnet was then fabricated and tested. The first test was made with the magnet immersed in liquid helium at 4.2K. The second test was conducted at 2K in vacuum. In the latter test, the magnet was conduction cooled from the mounting flange end.

  14. Inhibitor-neutralisation assay and electro-immuno assay of human factor IX (Christmas factor).

    PubMed

    Bertina, R M; van der Linden, I K

    1977-06-15

    A rabbit antibody specifically precipitating human factor IX has been used in the assay of factor IX antigen. The results obtained with two different methods (inhibitor-neutralisation assay and electro-immunoassay) have been compared in a group of healthy individuals and in a group of hemophilia B patients and carriers. In general, identical results are obtained with both methods, except in some hemophilia B+ carriers and patients, where the electroimmuno assay gives 1.5-2.0 times higher levels. Results obtained by electroimmuno assay are more accurate and reproducible than those obtained by inhibitor-neutralisation assay, which is of importance for its potential use in carrier detection.

  15. MAGNETIC IMAGING OF NANOCOMPOSITE MAGNETS

    SciTech Connect

    VOLKOV,V.V.ZHU,Y.

    2003-08-03

    Understanding the structure and magnetic behavior is crucial for optimization of nanocomposite magnets with high magnetic energy products. Many contributing factors such as phase composition, grain size distribution and specific domain configurations reflect a fine balance of magnetic energies at nanometer scale. For instance, magnetocrystalline anisotropy of grains and their orientations, degree of exchange coupling of magnetically soft and hard phases and specific energy of domain walls in a material. Modern microscopy, including Lorentz microscopy, is powerful tool for visualization and microstructure studies of nanocomposite magnets. However, direct interpretation of magnetically sensitive Fresnel/Foucault images for nanomagnets is usually problematic, if not impossible, because of the complex image contrast due to small grain size and sophisticated domain structure. Recently we developed an imaging technique based on Lorentz phase microscopy [l-4], which allows bypassing many of these problems and get quantitative information through magnetic flux mapping at nanometer scale resolution with a magnetically calibrated TEM [5]. This is our first report on application of this technique to nanocomposite magnets. In the present study we examine a nanocomposite magnet of nominal composition Nd{sub 2}Fe{sub 14+{delta}}B{sub 1.45} (14+{delta}=23.3, i.e. ''hard'' Nd{sub 2}Fe{sub 14}B-phase and 47.8 wt% of ''soft'' {alpha}-Fe phase ({delta}=9.3)), produced by Magnequench International, Inc. Conventional TEM/HREM study (Fig. 1-2) suggests that material has a bimodal grain-size distribution with maximum at d{sub max}=25 nm for Nd{sub 2}Fe{sub 14}B phase and d{sub max} = 15 nm for {alpha}-Fe phase (Fig.1c, Fig.2) in agreement with synchrotron X-ray studies (d{sub max}=23.5 nm for Nd{sub 2}Fe{sub 14}B [6]). Lattice parameters for Nd{sub 2}Fe{sub 14}B phase are a=8.80 and c=12.2 {angstrom}, as derived from SAED ring patterns (Fig.1a), again in good agreement with X-ray data

  16. Nondestructive assay confirmatory assessment experiments: mixed oxide

    SciTech Connect

    Lemming, J.F.

    1980-04-30

    The confirmatory assessment experiments demonstrate traceable nondestructive assay (NDA) measurements of plutonium in mixed oxide powder using commercially available spontaneous-fission assay systems. The experiments illustrate two major concepts: the production of calibration materials using calorimetric assay, and the use of paired measurements for measurement assurance. Two batches of well-characterized mixed oxide powder were used to establish the random and systematic error components. The major components of an NDA measurement assurance technique to establish and maintain traceability are identified and their functions are demonstrated. 20 refs., 10 figs., 10 tabs.

  17. Methods for threshold determination in multiplexed assays

    SciTech Connect

    Tammero, Lance F. Bentley; Dzenitis, John M; Hindson, Benjamin J

    2014-06-24

    Methods for determination of threshold values of signatures comprised in an assay are described. Each signature enables detection of a target. The methods determine a probability density function of negative samples and a corresponding false positive rate curve. A false positive criterion is established and a threshold for that signature is determined as a point at which the false positive rate curve intersects the false positive criterion. A method for quantitative analysis and interpretation of assay results together with a method for determination of a desired limit of detection of a signature in an assay are also described.

  18. Acoustic trapping as a generic non-contact incubation site for multiplex bead-based assays.

    PubMed

    Tenje, Maria; Xia, Hongyan; Evander, Mikael; Hammarström, Björn; Tojo, Axel; Belák, Sándor; Laurell, Thomas; LeBlanc, Neil

    2015-01-01

    In this study, we show a significantly reduced assay time and a greatly increased bead recovery for a commercial Luminex-based multiplex diagnostic immunoassay by performing all liquid handling steps of the assay protocol in a non-contact acoustic trapping platform. The Luminex assay is designed for detecting antibodies in poultry serum for infectious bursal disease virus, infectious bronchitis virus, Newcastle disease virus and avian reovirus. Here, we show proof-of-concept of a microfluidic system capable of being fully automated and handling samples in a parallel format with a miniature physical footprint where the affinity beads are retained in a non-contact levitated mode in a glass capillary throughout the assay protocol. The different steps are: incubation with the serum sample, secondary antibodies and fluorescent reporters and finally washing to remove any non-specifically bound species. A Luminex 200 instrument was used for the readout. The flow rates applied to the capillary during the initial trapping event and the wash steps were optimised for maximum bead recovery, resulting in a bead recovery of 75% for the complete assay. This can be compared to a bead recovery of approximately 30% when an automatic wash station was used when the assay was performed in the conventional manual format. The time for the incubation steps for a single assay was reduced by more than 50%, without affecting assay performance, since intermediate wash steps became redundant in the continuously perfused bead trapping capillary. We analyzed seven samples, in triplicates, and we can show that the readout of the assay performed in the acoustic trap compared 100% to the control ELISAs (positive or negative readout) and resulted in comparable S/P values as the conventional manual protocol. As the acoustic trapping does not require the particles to have magnetic properties, a greater degree of freedom in selecting microparticles can be provided. In extension, this can provide an

  19. Magnetism. Blowing magnetic skyrmion bubbles.

    PubMed

    Jiang, Wanjun; Upadhyaya, Pramey; Zhang, Wei; Yu, Guoqiang; Jungfleisch, M Benjamin; Fradin, Frank Y; Pearson, John E; Tserkovnyak, Yaroslav; Wang, Kang L; Heinonen, Olle; te Velthuis, Suzanne G E; Hoffmann, Axel

    2015-07-17

    The formation of soap bubbles from thin films is accompanied by topological transitions. Here we show how a magnetic topological structure, a skyrmion bubble, can be generated in a solid-state system in a similar manner. Using an inhomogeneous in-plane current in a system with broken inversion symmetry, we experimentally "blow" magnetic skyrmion bubbles from a geometrical constriction. The presence of a spatially divergent spin-orbit torque gives rise to instabilities of the magnetic domain structures that are reminiscent of Rayleigh-Plateau instabilities in fluid flows. We determine a phase diagram for skyrmion formation and reveal the efficient manipulation of these dynamically created skyrmions, including depinning and motion. The demonstrated current-driven transformation from stripe domains to magnetic skyrmion bubbles could lead to progress in skyrmion-based spintronics. PMID:26067256

  20. Magnetism. Blowing magnetic skyrmion bubbles.

    PubMed

    Jiang, Wanjun; Upadhyaya, Pramey; Zhang, Wei; Yu, Guoqiang; Jungfleisch, M Benjamin; Fradin, Frank Y; Pearson, John E; Tserkovnyak, Yaroslav; Wang, Kang L; Heinonen, Olle; te Velthuis, Suzanne G E; Hoffmann, Axel

    2015-07-17

    The formation of soap bubbles from thin films is accompanied by topological transitions. Here we show how a magnetic topological structure, a skyrmion bubble, can be generated in a solid-state system in a similar manner. Using an inhomogeneous in-plane current in a system with broken inversion symmetry, we experimentally "blow" magnetic skyrmion bubbles from a geometrical constriction. The presence of a spatially divergent spin-orbit torque gives rise to instabilities of the magnetic domain structures that are reminiscent of Rayleigh-Plateau instabilities in fluid flows. We determine a phase diagram for skyrmion formation and reveal the efficient manipulation of these dynamically created skyrmions, including depinning and motion. The demonstrated current-driven transformation from stripe domains to magnetic skyrmion bubbles could lead to progress in skyrmion-based spintronics.

  1. Human somatic mutation assays as biomarkers of carcinogenesis.

    PubMed Central

    Compton, P J; Hooper, K; Smith, M T

    1991-01-01

    This paper describes four assays that detect somatic gene mutations in humans: the hypoxanthine-guanine phosphoribosyl transferase assay, the glycophorin A assay, the HLA-A assay, and the sickle cell hemoglobin assay. Somatic gene mutation can be considered a biomarker of carcinogenesis, and assays for somatic mutation may assist epidemiologists in studies that attempt to identify factors associated with increased risks of cancer. Practical aspects of the use of these assays are discussed. PMID:1954924

  2. Human somatic mutation assays as biomarkers of carcinogenesis

    SciTech Connect

    Compton, P.J.E.; Smith, M.T. ); Hooper, K. )

    1991-08-01

    This paper describes four assays that detect somatic gene mutations in humans: the hypoxanthine-guanine phosphoribosyl transferase assay, the glycophorin A assay, the HLA-A assay, and the sickle cell hemoglobin assay. Somatic gene mutations can be considered a biomarker of carcinogenesis, and assays for somatic mutation may assist epidemiologists in studies that attempt to identify factors associated with increased risks of cancer. Practical aspects of the use of these assays are discussed.

  3. Proximity assays for sensitive quantification of proteins.

    PubMed

    Greenwood, Christina; Ruff, David; Kirvell, Sara; Johnson, Gemma; Dhillon, Harvinder S; Bustin, Stephen A

    2015-06-01

    Proximity assays are immunohistochemical tools that utilise two or more DNA-tagged aptamers or antibodies binding in close proximity to the same protein or protein complex. Amplification by PCR or isothermal methods and hybridisation of a labelled probe to its DNA target generates a signal that enables sensitive and robust detection of proteins, protein modifications or protein-protein interactions. Assays can be carried out in homogeneous or solid phase formats and in situ assays can visualise single protein molecules or complexes with high spatial accuracy. These properties highlight the potential of proximity assays in research, diagnostic, pharmacological and many other applications that require sensitive, specific and accurate assessments of protein expression. PMID:27077033

  4. Proximity assays for sensitive quantification of proteins

    PubMed Central

    Greenwood, Christina; Ruff, David; Kirvell, Sara; Johnson, Gemma; Dhillon, Harvinder S.; Bustin, Stephen A.

    2015-01-01

    Proximity assays are immunohistochemical tools that utilise two or more DNA-tagged aptamers or antibodies binding in close proximity to the same protein or protein complex. Amplification by PCR or isothermal methods and hybridisation of a labelled probe to its DNA target generates a signal that enables sensitive and robust detection of proteins, protein modifications or protein–protein interactions. Assays can be carried out in homogeneous or solid phase formats and in situ assays can visualise single protein molecules or complexes with high spatial accuracy. These properties highlight the potential of proximity assays in research, diagnostic, pharmacological and many other applications that require sensitive, specific and accurate assessments of protein expression. PMID:27077033

  5. BIOMARKER ASSAYS IN NIPPLE APIRATE FLUID

    EPA Science Inventory

    ABSTRACT

    The noninvasive technique of nipple aspiration as a potential source of biomarkers of breast cancer risk was evaluated. The feasibility of performing mutagenesis assays, amplifying DNA and performing protein electrophoresis on nipple aspirate fluid was explored. ...

  6. Electrochemical Assay of Gold-Plating Solutions

    NASA Technical Reports Server (NTRS)

    Chiodo, R.

    1982-01-01

    Gold content of plating solution is assayed by simple method that required only ordinary electrochemical laboratory equipment and materials. Technique involves electrodeposition of gold from solution onto electrode, the weight gain of which is measured. Suitable fast assay methods are economically and practically necessary in electronics and decorative-plating industries. If gold content in plating bath is too low, poor plating may result, with consequent economic loss to user.

  7. The comet assay: a heavenly method!

    PubMed

    Collins, Andrew R

    2015-01-01

    The contributions to this special issue of Mutagenesis have been selected to cover the main research areas served by the comet assay, namely genotoxicology, environmental toxicology, human biomonitoring and fundamental investigations into mechanisms of DNA damage and repair. Innovative methods are described, technical issues are explored, and guidelines are given for venturing into relatively new or unexploited areas of research. The popularity of the comet assay in a historical context is illustrated by a bibliometric survey.

  8. The comet assay in marine animals.

    PubMed

    Frenzilli, Giada; Lyons, Brett P

    2013-01-01

    Comet assay is a quick and versatile technique for assessing DNA damage in individual cells. It allows the detection of DNA single- and double-strand breaks, as well as the presence of alkali-labile sites and cross-links. Here we describe the protocols for the single-cell gel electrophoresis (Comet) assay in its alkaline (pH > 13), mild alkaline (pH = 12.1), and neutral (pH = 8) versions, when applied in marine animals.

  9. Automated optical sensing system for biochemical assays

    NASA Astrophysics Data System (ADS)

    Oroszlan, Peter; Duveneck, Gert L.; Ehrat, Markus; Widmer, H. M.

    1994-03-01

    In this paper, we present a new system called FOBIA that was developed and optimized with respect to automated operation of repetitive assay cycles with regenerable bioaffinity sensors. The reliability and precision of the new system is demonstrated by an application in a competitive assay for the detection of the triazine herbicide Atrazine. Using one sensor in more than 300 repetitive cycles, a signal precision better than 5% was achieved.

  10. Magnetic neurosurgery.

    PubMed

    Howard, M A; Grady, M S; Ritter, R C; Gillies, G T; Broaddus, W C; Dacey, R G

    1996-01-01

    Because of the complex shape of many brain structures, the ideal brain probe would be highly flexible and give the neurosurgeon the ability to independently and precisely control movement of the probe tip. A magnetic surgery system has been developed that implements this concept. Flexible brain probes with small permanent magnetic tips are placed on the brain surface through a small burr hole and then magnetically manipulated within the brain. Drive forces are produced by an array of six superconducting magnets suspended within a cryostatic helmet. They produce a maximum force of 3 times the threshold needed to move the probe through the parenchymal tissues at its normal speed of 1 mm/s. Computer-controlled magnetic impulses precisely direct the probe movement in any direction desired with movement accuracy of 0.47 mm in phantom gels. Probe position is monitored 3 times per second with orthogonally oriented microchannel plate X-ray systems, X-ray dose from a 3-hour simulated procedure is comparable to that of a chest X-ray. In vivo and in vitro feasibility studies have been carried out in dog and pig brains. Preclinical trials are planned for clinical applications including implantation of flexible brachytherapy threads.

  11. Protein immobilization techniques for microfluidic assays

    PubMed Central

    Kim, Dohyun; Herr, Amy E.

    2013-01-01

    Microfluidic systems have shown unequivocal performance improvements over conventional bench-top assays across a range of performance metrics. For example, specific advances have been made in reagent consumption, throughput, integration of multiple assay steps, assay automation, and multiplexing capability. For heterogeneous systems, controlled immobilization of reactants is essential for reliable, sensitive detection of analytes. In most cases, protein immobilization densities are maximized, while native activity and conformation are maintained. Immobilization methods and chemistries vary significantly depending on immobilization surface, protein properties, and specific assay goals. In this review, we present trade-offs considerations for common immobilization surface materials. We overview immobilization methods and chemistries, and discuss studies exemplar of key approaches—here with a specific emphasis on immunoassays and enzymatic reactors. Recent “smart immobilization” methods including the use of light, electrochemical, thermal, and chemical stimuli to attach and detach proteins on demand with precise spatial control are highlighted. Spatially encoded protein immobilization using DNA hybridization for multiplexed assays and reversible protein immobilization surfaces for repeatable assay are introduced as immobilization methods. We also describe multifunctional surface coatings that can perform tasks that were, until recently, relegated to multiple functional coatings. We consider the microfluidics literature from 1997 to present and close with a perspective on future approaches to protein immobilization. PMID:24003344

  12. Quantitation of flaviviruses by fluorescent focus assay.

    PubMed

    Payne, Anne F; Binduga-Gajewska, Iwona; Kauffman, Elizabeth B; Kramer, Laura D

    2006-06-01

    An indirect immunofluorescence assay for quantitation of flaviviruses was developed as an alternative to the standard plaque assay. The assay was validated with West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and Dengue virus (DENV) types 1-4. Vero cells were plated in 8-well chamber slides, and infected with 10-fold serial dilutions of virus. About 1-3 days after infection, cells were fixed, incubated with specific monoclonal antibody, and stained with a secondary antibody labeled with a fluorescent tag. Fluorescent foci of infection were observed and counted using a fluorescence microscope, and viral titers were calculated as fluorescent focus units (FFU) per ml. The optimal time for performing the fluorescent focus assay (FFA) on Vero cells was 24 h for WNV, and 48 h for SLEV and the four DENV serotypes. In contrast, the time required to complete a standard Vero cell plaque assay for these viruses range from 3 days for WNV to 11 days for DENV-1. Thus, the FFA method of virus titration is useful for viruses whose plaques develop slowly. In addition, these viruses can be quantitated by FFA on a mosquito cell line (C6/36), which does not support plaque formation. The FFA for flaviviruses was validated for accuracy, precision, specificity, and robustness of the assay.

  13. Scintillation Proximity Assay of Arginine Methylation

    PubMed Central

    Wu, Jiang; Xie, Nan; Feng, You; Zheng, Y. George

    2011-01-01

    Methylation of arginine residues, catalyzed by protein arginine methyltransferases (PRMTs), is one important protein post-translational modification involved in epigenetic regulation of gene expression. A fast and effective assay for PRMT can provide valuable information for dissecting the biological functions of PRMTs, as well as for screening small-molecule inhibitors of arginine methylation. Currently, among the methods used for PRMT activity measurement, many contain laborious separation procedures, which restrict the applications of these assays for high-throughput screening (HTS) in drug discovery. The authors report here a mix-and-measure method to measure PRMT activity based on the principle of scintillation proximity assay (SPA). In this assay, 3H-AdoMet was used as methyl donor, and biotin-modified histone H4 peptide served as a methylation substrate. Following the methylation reaction catalyzed by PRMTs, streptavidin-coated SPA beads were added to the reaction solution, and SPA signals were detected by a MicroBeta scintillation counter. No separation step is needed, which simplifies the assay procedure and greatly enhances the assay speed. Particularly, the miniaturization and robustness suggest that this method is suited for HTS of PRMT inhibitors. PMID:21821785

  14. Dot-immunobinding assay (Dot-Iba).

    PubMed

    Surendran, Sumi; Mathai, Annamma; Radhakrishnan, Vishnampet Venkataraman

    2015-01-01

    Dot-immunobinding assay (Dot-Iba) is a simple and highly reproducible immunodiagnostic method. Antibody or antigen is dotted directly onto nitrocellulose membrane (NCM) discs. The diagnostic material to be checked can be incubated on this disc. Presence of antigen-antibody complex in NCM discs can be directly demonstrated with enzyme-conjugated antiglobulins and substrate. Development of a purple-pink colored, insoluble substrate product in the nitrocellulose membrane will be considered a positive result in the assay. This assay allows the processing of multiple specimens at a time and the entire operational procedures required only 4-6 h. Dot-IBA is rapid and the technical steps involved in the assay are much simpler than the other immunoassays such as enzyme-linked immunosorbent assay in detecting circulating antigen and antibody in clinical samples. The Dot-Iba showed an overall sensitivity of 60 % for tuberculous meningitis diagnosis and no false positive results were encountered. Hence this assay is highly specific for the diagnosis of paucibacillary diseases like extrapulmonary tuberculosis. Dot-Iba is best suited to laboratories in developing world where there are constraints in laboratory resources.

  15. Radioimmune assay of human platelet prostaglandin synthetase

    SciTech Connect

    Roth, G.J.; Machuga, E.T.

    1982-02-01

    Normal platelet function depends, in part, on platelet PG synthesis. PG synthetase (cyclo-oxygenase) catalyzes the first step in PG synthesis, the formation of PGH/sub 2/ from arachidonic acid. Inhibition of the enzyme by ASA results in an abnormality in the platelet release reaction. Patients with pparent congenital abnormalities in the enzyme have been described, and the effects have been referred to as ''aspirin-like'' defects of the platelet function. These patients lack platelet PG synthetase activity, but the actual content of PG synthetase protein in these individuals' platelets is unknown. Therefore an RIA for human platelet PG synthetase would provide new information, useful in assessing the aspirin-like defects of platelet function. An RIA for human platelet PG synthetase is described. The assay utilizes a rabbit antibody directed against the enzyme and (/sup 125/I)-labelled sheep PG synthetase as antigen. The human platelet enzyme is assayed by its ability to inhibit precipitation of the (/sup 125/I)antigen. The assay is sensitive to 1 ng of enzyme. By the immune assay, human platelets contain approximately 1200 ng of PG synethetase protein per 1.5 mg of platelet protein (approximately 10/sup 9/ platelets). This content corresponds to 10,000 enzyme molecules per platelet. The assay provides a rapid and convenient assay for the human platelet enzyme, and it can be applied to the assessment of patients with apparent platelet PG synthetase (cyclo-oxygenase) deficiency.

  16. Synthesis and characterization of functionalized magnetic nanoparticles

    NASA Astrophysics Data System (ADS)

    Biswal, Dipti; Peeples, Brianna N.; Spence, Destiny D.; Peeples, Caryn; Bell, Crystal N.; Pradhan, A. K.

    2012-04-01

    Magnetic nanoparticles have been used in a wide array of industrial and biomedical applications due to their unique properties at the nanoscale level. They are extensively used in magnetic resonance imaging (MRI), magnetic hyperthermia treatment, drug delivery, and in assays for biological separations. Furthermore, superparamagnetic nanoparticles are of large interest for in vivo applications. However, these unmodified nanoparticles aggregate and consequently lose their superparamagnetic behaviors, due to high surface to volume ratio and strong dipole to dipole interaction. For these reasons, surface coating is necessary for the enhancement and effectiveness of magnetic nanoparticles to be used in various applications. In addition to providing increased stability to the nanoparticles in different solvents or media, stabilizers such as surfactants, organic/inorganic molecules, polymer and co-polymers are employed as surface coatings, which yield magnetically responsive systems. In this work we present the synthesis and magnetic characterization of Fe3O4 nanoparticles coated with 3-aminopropyltriethoxy silane (APS) and citric acid. The particles magnetic hysteresis was measured by a superconducting quantum interference device (SQUID) magnetometer with an in-plane magnetic field. The uncoated and coated magnetic nanoparticles were characterized by using fourier transform infrared (FTIR), UV-vis, X-ray diffraction, transmission electron microscopy, and thermo-gravimetric analysis.

  17. Bead-Based Assays for Biodetection: From Flow-Cytometry to Microfluidics

    SciTech Connect

    Ozanich, Richard M.; Antolick, Kathryn C.; Bruckner-Lea, Cindy J.; Bunch, Kyle J.; Dockendorff, Brian P.; Grate, Jay W.; Nash, Michael A.; Tyler, Abby J.

    2009-05-04

    ABSTRACT The potential for the use of biological agents by terrorists is a real threat. Two approaches for detection of biological species will be described: 1) The use of microbead arrays for multiplexed flow cytometry detection of cytokines and botulinum neurotoxin simulant, and 2) a microfluidic platform for capture and separation of different size superparamagnetic nanoparticles followed by on-chip fluorescence detection of the sandwich complex. The methods and automated fluidic systems used for trapping functionalized microbeads will be described. This approach allows sample, assay reagents, and wash solutions to be perfused over a micro-column of beads, resulting in faster and more sensitive assays. The automated fluidic approach resulted in up to five-fold improvements in assay sensitivity/speed as compared to identical assays performed in a typical manual batch mode. A second approach for implementing multiplexed bead-based assays without using flow cytometry detection is currently under development. The goal of the microfluidic-based approach is to achieve rapid (<20 minutes), multiplexed (> 3 bioagents) detection using a simple and low-cost, integrated microfluidic/optical detection platform. Using fiber-optic guided laser-induced fluorescence, assay detection limits were shown to be in the 100’s of picomolar range (10’s of micrograms per liter) for botulinum neurotoxin simulant without any optimization of the microfluidic device or optical detection approach. Video taping magnetic nanoparticle capture and release was used to improve understanding of the process and revealed interesting behavior.

  18. Magnetic light

    PubMed Central

    Kuznetsov, Arseniy I.; Miroshnichenko, Andrey E.; Fu, Yuan Hsing; Zhang, JingBo; Luk’yanchuk, Boris

    2012-01-01

    Spherical silicon nanoparticles with sizes of a few hundreds of nanometers represent a unique optical system. According to theoretical predictions based on Mie theory they can exhibit strong magnetic resonances in the visible spectral range. The basic mechanism of excitation of such modes inside the nanoparticles is very similar to that of split-ring resonators, but with one important difference that silicon nanoparticles have much smaller losses and are able to shift the magnetic resonance wavelength down to visible frequencies. We experimentally demonstrate for the first time that these nanoparticles have strong magnetic dipole resonance, which can be continuously tuned throughout the whole visible spectrum varying particle size and visually observed by means of dark-field optical microscopy. These optical systems open up new perspectives for fabrication of low-loss optical metamaterials and nanophotonic devices. PMID:22768382

  19. MAGNETIC GRID

    DOEpatents

    Post, R.F.

    1960-08-01

    An electronic grid is designed employing magnetic forces for controlling the passage of charged particles. The grid is particularly applicable to use in gas-filled tubes such as ignitrons. thyratrons, etc., since the magnetic grid action is impartial to the polarity of the charged particles and, accordingly. the sheath effects encountered with electrostatic grids are not present. The grid comprises a conductor having sections spaced apart and extending in substantially opposite directions in the same plane, the ends of the conductor being adapted for connection to a current source.

  20. Magnetic particles

    NASA Technical Reports Server (NTRS)

    Chang, Manchium (Inventor); Colvin, Michael S. (Inventor)

    1989-01-01

    Magnetic polymer particles are formed by swelling porous, polymer particles and impregnating the particles with an aqueous solution of precursor magnetic metal salt such as an equimolar mixture of ferrous chloride and ferric chloride. On addition of a basic reagent such as dilute sodium hydroxide, the metal salts are converted to crystals of magnetite which are uniformly contained througout the pores of the polymer particle. The magnetite content can be increased and neutral buoyancy achieved by repetition of the impregnaton and neutralization steps to adjust the magnetite content to a desired level.

  1. Controlling variation in the comet assay

    PubMed Central

    Collins, Andrew R.; El Yamani, Naouale; Lorenzo, Yolanda; Shaposhnikov, Sergey; Brunborg, Gunnar; Azqueta, Amaya

    2014-01-01

    Variability of the comet assay is a serious issue, whether it occurs from experiment to experiment in the same laboratory, or between different laboratories analysing identical samples. Do we have to live with high variability, just because the comet assay is a biological assay rather than analytical chemistry? Numerous attempts have been made to limit variability by standardizing the assay protocol, and the critical steps in the assay have been identified; agarose concentration, duration of alkaline incubation, and electrophoresis conditions (time, temperature, and voltage gradient) are particularly important. Even when these are controlled, variation seems to be inevitable. It is helpful to include in experiments reference standards, i.e., cells with a known amount of specific damage to the DNA. They can be aliquots frozen from a single large batch of cells, either untreated (negative controls) or treated with, for example, H2O2 or X-rays to induce strand breaks (positive control for the basic assay), or photosensitiser plus light to oxidize guanine (positive control for Fpg- or OGG1-sensitive sites). Reference standards are especially valuable when performing a series of experiments over a long period—for example, analysing samples of white blood cells from a large human biomonitoring trial—to check that the assay is performing consistently, and to identify anomalous results necessitating a repeat experiment. The reference values of tail intensity can also be used to iron out small variations occurring from day to day. We present examples of the use of reference standards in human trials, both within one laboratory and between different laboratories, and describe procedures that can be used to control variation. PMID:25368630

  2. Cell Culture Assay for Human Noroviruses [response

    SciTech Connect

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  3. Measurement of dabigatran in standardly used clinical assays, whole blood viscoelastic coagulation, and thrombin generation assays.

    PubMed

    van Ryn, Joanne; Grottke, Oliver; Spronk, Henri

    2014-09-01

    Dabigatran, a direct thrombin inhibitor, is increasingly used clinically as one of the new oral anticoagulants. This review summarizes the assays available to measure its activity and includes the relative sensitivity of the different assays for this agent. In addition to plasma-based clotting tests, assays commonly used in surgical/emergency settings, such as activated clotting time and thromboelastometry/thromboelastography, are reviewed. In addition, the thrombin generation assay is discussed as an important method to determine the potential risk of thrombosis or bleeding and its relevance to the measurement of direct thrombin inhibitors. PMID:25168938

  4. Using the CPTAC Assay Portal to Identify and Implement Highly Characterized Targeted Proteomics Assays.

    PubMed

    Whiteaker, Jeffrey R; Halusa, Goran N; Hoofnagle, Andrew N; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John A; Kennedy, Jacob; Mani, D R; Zimmerman, Lisa J; Meyer, Matthew R; Mesri, Mehdi; Boja, Emily; Carr, Steven A; Chan, Daniel W; Chen, Xian; Chen, Jing; Davies, Sherri R; Ellis, Matthew J C; Fenyö, David; Hiltke, Tara; Ketchum, Karen A; Kinsinger, Chris; Kuhn, Eric; Liebler, Daniel C; Liu, Tao; Loss, Michael; MacCoss, Michael J; Qian, Wei-Jun; Rivers, Robert; Rodland, Karin D; Ruggles, Kelly V; Scott, Mitchell G; Smith, Richard D; Thomas, Stefani; Townsend, R Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Rodriguez, Henry; Paulovich, Amanda G

    2016-01-01

    The Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal (http://assays.cancer.gov) to serve as an open-source repository of well-characterized targeted proteomic assays. The portal is designed to curate and disseminate highly characterized, targeted mass spectrometry (MS)-based assays by providing detailed assay performance characterization data, standard operating procedures, and access to reagents. Assay content is accessed via the portal through queries to find assays targeting proteins associated with specific cellular pathways, protein complexes, or specific chromosomal regions. The position of the peptide analytes for which there are available assays are mapped relative to other features of interest in the protein, such as sequence domains, isoforms, single nucleotide polymorphisms, and posttranslational modifications. The overarching goals are to enable robust quantification of all human proteins and to standardize the quantification of targeted MS-based assays to ultimately enable harmonization of results over time and across laboratories. PMID:26867747

  5. Using the CPTAC Assay Portal to identify and implement highly characterized targeted proteomics assays

    PubMed Central

    Whiteaker, Jeffrey R; Halusa, Goran N; Hoofnagle, Andrew N; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John A; Kennedy, Jacob; Mani, DR; Zimmerman, Lisa J; Meyer, Matthew R.; Mesri, Mehdi; Abbatiello, Susan E; Boja, Emily; Carr, Steven A.; Chan, Daniel W.; Chen, Xian; Chen, Jing; Davies, Sherri R; Ellis, Matthew J. C.; Fenyö, David; Hiltke, Tara; Ketchum, Karen A.; Kinsinger, Chris; Kuhn, Eric; Liebler, Daniel C.; Lin, De; Liu, Tao; Loss, Michael; MacCoss, Michael J; Qian, Wei-Jun; Rivers, Robert; Rodland, Karin D.; Ruggles, Kelly V; Scott, Mitchell G; Smith, Richard D.; Thomas, Stefani; Townsend, R. Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Rodriguez, Henry; Paulovich, Amanda G

    2016-01-01

    Summary The Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal (http://assays.cancer.gov) to serve as an open-source repository of well-characterized targeted proteomic assays. The portal is designed to curate and disseminate highly characterized, targeted mass spectrometry (MS)-based assays by providing detailed assay performance characterization data, standard operating procedures, and access to reagents. Assay content is accessed via the portal through queries to find assays targeting proteins associated with specific cellular pathways, protein complexes, or specific chromosomal regions. The position of the peptide analytes for which there are available assays are mapped relative to other features of interest in the protein, such as sequence domains, isoforms, single nucleotide polymorphisms, and post-translational modifications. The overarching goals are to enable robust quantification of all human proteins and to standardize the quantification of targeted MS-based assays to ultimately enable harmonization of results over time and across laboratories. PMID:26867747

  6. Superconducting Magnets for Magnetic Resonance Imaging

    NASA Astrophysics Data System (ADS)

    Feenan, Peter

    2000-03-01

    MRI is now a well established diagnostic technique in medicine. The richness of information provided by magnetic resonance gives rise to a variety of techniques which in turn leads to a variety of magnet designs. Magnet designers must consider suitable superconduting materials for the magnet, but need also to consider the overall fomat of the magnet to maximise patient comfort, access for clinicians and convenience of use - in some examples magnets are destined for use within the operating theatre and special considerations are required for this. Magnet types include; (1) low-field general purpose imagers, (2) extremity imaging, (3) open magnets with exellent all-round access often employing iron or permanent magnetic materials, (4) high-field magnets, and (5) very high-field (7 Tesla and more) magnets for spectroscopy and functional imaging research. Examples of these magnet varieties will be shown and some of the design challenges discussed.

  7. Magnetic tape

    NASA Technical Reports Server (NTRS)

    Robinson, Harriss

    1992-01-01

    The move to visualization and image processing in data systems is increasing the demand for larger and faster mass storage systems. The technology of choice is magnetic tape. This paper briefly reviews the technology past, present, and projected. A case is made for standards and the value of the standards to users.

  8. Magnetic Liquids

    NASA Technical Reports Server (NTRS)

    1987-01-01

    Ferrofluidics Corporation's recent innovation is a spindle for rotating computer discs that supports the disc's rotating shaft on a film of magnetic fluid instead of conventional ball bearings. According to its developers, the spindle offers greatly increased rotational stability, meaning substantially reduced vibration and mechanical noise, and non- repeatable runout. This allows disc drives to store two to 10 times more information.

  9. Research highlights: digital assays on chip.

    PubMed

    Kim, Donghyuk; Wei, Qingshan; Kong, Janay Elise; Ozcan, Aydogan; Di Carlo, Dino

    2015-01-01

    The ability to break up a volume of fluid into smaller pieces that are confined or separated to prevent molecular communication/transport is a key capability intrinsic to microfluidic systems. This capability has been used to develop or implement digital versions of traditional molecular analysis assays, including digital PCR and digital immunoassays/ELISA. In these digital versions, the concentration of the target analyte is in a range such that, when sampled into smaller fluid volumes, either a single molecule or no molecule may be present. Subsequent amplification is sensitive enough to obtain a digital readout of the presence of these target molecules. Advantages of such approaches that are claimed include quantification without calibration and robustness to variations in reaction conditions or times because the digital readout is less sensitive to absolute signal intensity levels. Weaknesses of digital approaches include a lower dynamic range of concentrations over which the assay is sensitive, which depends on the total volume that can be analyzed. We highlight recent efforts to expand the dynamic range of digital assays based on exploiting reaction/diffusion phenomena. A side-by-side study that evaluates the strengths of digital assays reveals that the majority of these claims are supported, with specific caveats. Finally, we highlight approaches to apply digital assays to analyze new types of reactions, including the active transport of protons across membranes by ATPases at the single protein level - perhaps opening up new biophysical understanding and screening opportunities, similar to widely deployed single-molecule ion channel analysis.

  10. Multi-platform metabolomics assays for human lung lavage fluids in an air pollution exposure study.

    PubMed

    Surowiec, Izabella; Karimpour, Masoumeh; Gouveia-Figueira, Sandra; Wu, Junfang; Unosson, Jon; Bosson, Jenny A; Blomberg, Anders; Pourazar, Jamshid; Sandström, Thomas; Behndig, Annelie F; Trygg, Johan; Nording, Malin L

    2016-07-01

    Metabolomics protocols are used to comprehensively characterize the metabolite content of biological samples by exploiting cutting-edge analytical platforms, such as gas chromatography (GC) or liquid chromatography (LC) coupled to mass spectrometry (MS) assays, as well as nuclear magnetic resonance (NMR) assays. We have developed novel sample preparation procedures combined with GC-MS, LC-MS, and NMR metabolomics profiling for analyzing bronchial wash (BW) and bronchoalveolar lavage (BAL) fluid from 15 healthy volunteers following exposure to biodiesel exhaust and filtered air. Our aim was to investigate the responsiveness of metabolite profiles in the human lung to air pollution exposure derived from combustion of biofuels, such as rapeseed methyl ester biodiesel, which are increasingly being promoted as alternatives to conventional fossil fuels. Our multi-platform approach enabled us to detect the greatest number of unique metabolites yet reported in BW and BAL fluid (82 in total). All of the metabolomics assays indicated that the metabolite profiles of the BW and BAL fluids differed appreciably, with 46 metabolites showing significantly different levels in the corresponding lung compartments. Furthermore, the GC-MS assay revealed an effect of biodiesel exhaust exposure on the levels of 1-monostearylglycerol, sucrose, inosine, nonanoic acid, and ethanolamine (in BAL) and pentadecanoic acid (in BW), whereas the LC-MS assay indicated a shift in the levels of niacinamide (in BAL). The NMR assay only identified lactic acid (in BW) as being responsive to biodiesel exhaust exposure. Our findings demonstrate that the proposed multi-platform approach is useful for wide metabolomics screening of BW and BAL fluids and can facilitate elucidation of metabolites responsive to biodiesel exhaust exposure. Graphical Abstract Graphical abstract illustrating the study workflow. NMR Nuclear Magnetic Resonance, LC-TOFMS Liquid chromatography-Time Of Flight Mass Spectrometry, GC Gas

  11. Competitive protein binding assay for piritrexim

    SciTech Connect

    Woolley, J.L. Jr.; Ringstad, J.L.; Sigel, C.W. )

    1989-09-01

    A competitive protein binding assay for piritrexim (PTX, 1) that makes use of a commercially available radioassay kit for methotrexate has been developed. After it is selectively extracted from plasma, PTX competes with ({sup 125}I)methotrexate for binding to dihydrofolate reductase isolated from Lactobacillus casei. Free drug is separated from bound drug by adsorption to dextran-coated charcoal. Piritrexim is measurable over a range of 0.01 to 10.0 micrograms/mL in plasma with a coefficient of variation less than 15%. The limit of sensitivity of the assay is approximately 2 ng/mL. An excellent correlation between this assay and a previously published HPLC method was found.

  12. Nuclear Resonance Fluorescence for Materials Assay

    SciTech Connect

    Quiter, Brian; Ludewigt, Bernhard; Mozin, Vladimir; Prussin, Stanley

    2009-06-05

    This paper discusses the use of nuclear resonance fluorescence (NRF) techniques for the isotopic and quantitative assaying of radioactive material. Potential applications include age-dating of an unknown radioactive source, pre- and post-detonation nuclear forensics, and safeguards for nuclear fuel cycles Examples of age-dating a strong radioactive source and assaying a spent fuel pin are discussed. The modeling work has ben performed with the Monte Carlo radiation transport computer code MCNPX, and the capability to simulate NRF has bee added to the code. Discussed are the limitations in MCNPX's photon transport physics for accurately describing photon scattering processes that are important contributions to the background and impact the applicability of the NRF assay technique.

  13. Nuclear Resonance Fluorescence for Materials Assay

    SciTech Connect

    Quiter, Brian J.; Ludewigt, Bernhard; Mozin, Vladimir; Prussin, Stanley

    2009-06-29

    This paper discusses the use of nuclear resonance fluorescence (NRF) techniques for the isotopic and quantitative assaying of radioactive material. Potential applications include age-dating of an unknown radioactive source, pre- and post-detonation nuclear forensics, and safeguards for nuclear fuel cycles Examples of age-dating a strong radioactive source and assaying a spent fuel pin are discussed. The modeling work has ben performed with the Monte Carlo radiation transport computer code MCNPX, and the capability to simulate NRF has bee added to the code. Discussed are the limitations in MCNPX?s photon transport physics for accurately describing photon scattering processes that are important contributions to the background and impact the applicability of the NRF assay technique.

  14. Assays for aptamer-based platforms.

    PubMed

    Citartan, Marimuthu; Gopinath, Subash C B; Tominaga, Junji; Tan, Soo-Choon; Tang, Thean-Hock

    2012-04-15

    Aptamers are single stranded DNA or RNA oligonucleotides that have high affinity and specificity towards a wide range of target molecules. Aptamers have low molecular weight, amenable to chemical modifications and exhibit stability undeterred by repetitive denaturation and renaturation. Owing to these indispensable advantages, aptamers have been implemented as molecular recognition element as alternative to antibodies in various assays for diagnostics. By amalgamating with a number of methods that can provide information on the aptamer-target complex formation, aptamers have become the elemental tool for numerous biosensor developments. In this review, administration of aptamers in applications involving assays of fluorescence, electrochemistry, nano-label and nano-constructs are discussed. Although detection strategies are different for various aptamer-based assays, the core of the design strategies is similar towards reporting the presence of specific target binding to the corresponding aptamers. It is prognosticated that aptamers will find even broader applications with the development of new methods of transducing aptamer target binding.

  15. Miniaturization of hydrolase assays in thermocyclers.

    PubMed

    Lucena, Severino A; Moraes, Caroline S; Costa, Samara G; de Souza, Wanderley; Azambuja, Patrícia; Garcia, Eloi S; Genta, Fernando A

    2013-03-01

    We adapted the protocols of reducing sugar measurements with dinitrosalicylic acid and bicinchoninic acid for thermocyclers and their use in enzymatic assays for hydrolases such as amylase and β-1,3-glucanase. The use of thermocyclers for these enzymatic assays resulted in a 10 times reduction in the amount of reagent and volume of the sample needed when compared with conventional microplate protocols. We standardized absorbance readings from the polymerase chain reaction plates, which allowed us to make direct readings of the techniques above, and a β-glycosidase assay was also established under the same conditions. Standardization of the enzymatic reaction in thermocyclers resulted in less time-consuming temperature calibrations and without loss of volume through leakage or evaporation from the microplate. Kinetic parameters were successfully obtained, and the use of the thermocycler allowed the measurement of enzymatic activities in biological samples from the field with a limited amount of protein. PMID:23123426

  16. Flow cytometer measurement of binding assays

    DOEpatents

    Saunders, George C.

    1987-01-01

    A method of measuring the result of a binding assay that does not require separation of fluorescent smaller particles is disclosed. In a competitive binding assay the smaller fluorescent particles coated with antigen compete with antigen in the sample being analyzed for available binding sites on larger particles. In a sandwich assay, the smaller, fluorescent spheres coated with antibody attach themselves to molecules containing antigen that are attached to larger spheres coated with the same antibody. The separation of unattached, fluorescent smaller particles is made unnecessary by only counting the fluorescent events triggered by the laser of a flow cytometer when the event is caused by a particle with a light scatter measurement within a certain range corresponding to the presence of larger particles.

  17. Fungicide resistance assays for fungal plant pathogens.

    PubMed

    Secor, Gary A; Rivera, Viviana V

    2012-01-01

    Fungicide resistance assays are useful to determine if a fungal pathogen has developed resistance to a fungicide used to manage the disease it causes. Laboratory assays are used to determine loss of sensitivity, or resistance, to a fungicide and can explain fungicide failures and for developing successful fungicide recommendations in the field. Laboratory assays for fungicide resistance are conducted by measuring reductions in growth or spore germination of fungi in the presence of fungicide, or by molecular procedures. This chapter describes two techniques for measuring fungicide resistance, using the sugarbeet leaf spot fungus Cercospora beticola as a model for the protocol. Two procedures are described for fungicides from two different classes; growth reduction for triazole (sterol demethylation inhibitor; DMI) fungicides, and inhibition of spore germination for quinone outside inhibitor (QoI) fungicides.

  18. The Soft Agar Colony Formation Assay

    PubMed Central

    Borowicz, Stanley; Van Scoyk, Michelle; Avasarala, Sreedevi; Karuppusamy Rathinam, Manoj Kumar; Tauler, Jordi; Bikkavilli, Rama Kamesh; Winn, Robert A.

    2014-01-01

    Anchorage-independent growth is the ability of transformed cells to grow independently of a solid surface, and is a hallmark of carcinogenesis. The soft agar colony formation assay is a well-established method for characterizing this capability in vitro and is considered to be one of the most stringent tests for malignant transformation in cells. This assay also allows for semi-quantitative evaluation of this capability in response to various treatment conditions. Here, we will demonstrate the soft agar colony formation assay using a murine lung carcinoma cell line, CMT167, to demonstrate the tumor suppressive effects of two members of the Wnt signaling pathway, Wnt7A and Frizzled-9 (Fzd-9). Concurrent overexpression of Wnt7a and Fzd-9 caused an inhibition of colony formation in CMT167 cells. This shows that expression of Wnt7a ligand and its Frizzled-9 receptor is sufficient to suppress tumor growth in a murine lung carcinoma model. PMID:25408172

  19. Neutron Assay System for Confinement Vessel Disposition

    SciTech Connect

    Frame, Katherine C; Bourne, Mark M; Crooks, William J; Evans, Louise; Mayo, Douglas R; Miko, David K; Salazar, William R; Stange, Sy; Valdez, Jose I; Vigil, Georgiana M

    2012-07-13

    Waste will be removed from confinement vessels remaining from 1970s-era experiments. Los Alamos has 9+ spherical confinement vessels remaining from experiments. Each vessel contains {approx} 500 lbs of radioactive debris such as actinide metals and oxides, metals, powdered silica, graphite, and wires and hardware. In order to dispose of the vessels, debris and contamination must be removed. Neutron assay system was designed to assay vessels before and after cleanout. System requirements are: (1) Modular and moveable; (2) Capable of detecting {approx}100g {sup 239}Pu equivalent in a 2-inch thick steel sphere with 6 foot diameter; and (3) Capable of safeguards-quality assays. Initial design parameters arethe use of 4-atm {sup 3}He tubes with length of 6 feet, and {sup 3}He tubes embedded in polyethelene for moderation. This paper describes the calibration of the Confinement Vessel Assay System (CVAS) and quantification of its uncertainties. Assay uncertainty depends on five factors: (1) Statistical uncertainty in the assay measurement; (2) Statistical uncertainty in the background measurement; (3) Statistical uncertainty in the isotopics determination - This should be much smaller than the other uncertainties; (4) Systematic uncertainty due to position bias; and (5) Systematic uncertainty due to fluctuations in cosmic ray spallation. This one can be virtually eliminated by performing the background measurement with an empty vessel - but that may not be possible. We used modeling and experiments to quantify the systematic uncertainties. The calibration assumes a uniform distribution of material, but reality will be different. MCNPX modeling was used to quantify the positional bias. The model was benchmarked to build confidence in its results. Material at top of vessel is 44% greater than amount assayed, according to singles. Material near 19-tube detector is 38% less than amount assayed, according to singles. Cosmic ray spallation contributes significantly to the

  20. A new fluorescent assay for enalapril maleate.

    PubMed

    de los A Oliva, María; Sombra, Lorena L; Olsina, Roberto A; Masi, Adriana N

    2005-09-01

    A new spectrofluorimetric method for the enalapril maleate monitoring was studied. Enalapril maleate was found to be highly photolabile. This drug was evaluated according to photodegradation assay at pH 2.5 and 6. Enalapril maleate was exposed to UVA-UVB radiations. Under these specific conditions was found as degradation product, the diketopiperazine. The modification of the fluorescent properties of enalapril maleate in solution after exposure UV-radiation and the degradation mechanisms were studied. The photodegradation was followed by the developed spectrofluorimetric assay.

  1. Technique to optimize magnetic response of gelatin coated magnetic nanoparticles.

    PubMed

    Parikh, Nidhi; Parekh, Kinnari

    2015-07-01

    The paper describes the results of optimization of magnetic response for highly stable bio-functionalize magnetic nanoparticles dispersion. Concentration of gelatin during in situ co-precipitation synthesis was varied from 8, 23 and 48 mg/mL to optimize magnetic properties. This variation results in a change in crystallite size from 10.3 to 7.8 ± 0.1 nm. TEM measurement of G3 sample shows highly crystalline spherical nanoparticles with a mean diameter of 7.2 ± 0.2 nm and diameter distribution (σ) of 0.27. FTIR spectra shows a shift of 22 cm(-1) at C=O stretching with absence of N-H stretching confirming the chemical binding of gelatin on magnetic nanoparticles. The concept of lone pair electron of the amide group explains the mechanism of binding. TGA shows 32.8-25.2% weight loss at 350 °C temperature substantiating decomposition of chemically bind gelatin. The magnetic response shows that for 8 mg/mL concentration of gelatin, the initial susceptibility and saturation magnetization is the maximum. The cytotoxicity of G3 sample was assessed in Normal Rat Kidney Epithelial Cells (NRK Line) by MTT assay. Results show an increase in viability for all concentrations, the indicative probability of a stimulating action of these particles in the nontoxic range. This shows the potential of this technique for biological applications as the coated particles are (i) superparamagnetic (ii) highly stable in physiological media (iii) possibility of attaching other drug with free functional group of gelatin and (iv) non-toxic.

  2. Instructions for Uploading Data to the Assay Portal - Instructions for Uploading Data to the Assay Portal

    Cancer.gov

    This document provides instructions for configuring and uploading data files to the CPTAC Assay Portal. It is divided into sections, with an overview checklist provided at the end. If help is needed at any stage of the process, please use the support page: https://assays.cancer.gov/support/

  3. Magnetic bearing. [for supplying magnetic fluxes

    NASA Technical Reports Server (NTRS)

    Studer, P. A. (Inventor)

    1975-01-01

    A magnetic bearing is described which includes a pair of coaxial, toroidal, and permanent magnets having axially directed poles. Like poles of the permanent magnets are adjacent to each other, whereby the permanent magnets have a tendency to be urged apart along the common axis. An electromagnet is wound coaxially with the permanent magnets in such a manner that the poles are axially directed. Between the poles of each permanent magnet there is a low magnetic reluctance circuit including two series air gaps. Between the poles of the electromagnet a low reluctance path including only one air gap of each of the low magnetic reluctance circuits is provided. The low reluctance path for the electromagnet includes a ring axially translatable relative to the permanent magnets. The ring forms opposite faces of the air gaps in the magnetic circuits for each permanent magnet.

  4. Magnetic Reconnection

    SciTech Connect

    Masaaki Yamada, Russell Kulsrud and Hantao Ji

    2009-09-17

    We review the fundamental physics of magnetic reconnection in laboratory and space plasmas, by discussing results from theory, numerical simulations, observations from space satellites, and the recent results from laboratory plasma experiments. After a brief review of the well-known early work, we discuss representative recent experimental and theoretical work and attempt to interpret the essence of significant modern findings. In the area of local reconnection physics, many significant findings have been made with regard to two- uid physics and are related to the cause of fast reconnection. Profiles of the neutral sheet, Hall currents, and the effects of guide field, collisions, and micro-turbulence are discussed to understand the fundamental processes in a local reconnection layer both in space and laboratory plasmas. While the understanding of the global reconnection dynamics is less developed, notable findings have been made on this issue through detailed documentation of magnetic self-organization phenomena in fusion plasmas. Application of magnetic reconnection physics to astrophysical plasmas is also brie y discussed.

  5. Optical protein detection based on magnetic clusters rotation.

    PubMed

    Ramiandrisoa, Donatien; Brient-Litzler, Elodie; Daynes, Aurélien; Compain, Eric; Bibette, Jérôme; Baudry, Jean

    2015-09-25

    In this paper we present a simple method to quantify aggregates of 200nm magnetic particles. This method relies on the optical and magnetic anisotropy of particle aggregates, whereas dispersed particles are optically isotropic. We orientate aggregates by applying short pulses of a magnetic field, and we measure optical density variation directly linked to this reorientation. By computing the scattering efficiency of doublets and singlets, we demonstrate the absolute quantification of a few % of doublets in a well dispersed suspension. More generally, these optical variations are related to the aggregation state of the sample. This method can be easily applied to an agglutination assay, where target proteins induce aggregation of colloidal particles. By observing only aligned clusters, we increase sensitivity and we reduce the background noise as compared to a classical agglutination assay: we obtain a detection limit on the C-reactive protein of less than 3pM for a total assay time of 10min.

  6. 21 CFR 864.7425 - Carboxyhemoglobin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Carboxyhemoglobin assay. 864.7425 Section 864.7425 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7425...

  7. 21 CFR 864.7490 - Sulfhemoglobin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Sulfhemoglobin assay. 864.7490 Section 864.7490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7490...

  8. 21 CFR 864.7490 - Sulfhemoglobin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Sulfhemoglobin assay. 864.7490 Section 864.7490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7490...

  9. 21 CFR 864.7425 - Carboxyhemoglobin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Carboxyhemoglobin assay. 864.7425 Section 864.7425 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7425...

  10. 21 CFR 864.7425 - Carboxyhemoglobin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Carboxyhemoglobin assay. 864.7425 Section 864.7425 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7425...

  11. 21 CFR 864.7490 - Sulfhemoglobin assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Sulfhemoglobin assay. 864.7490 Section 864.7490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7490...

  12. 21 CFR 864.7490 - Sulfhemoglobin assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Sulfhemoglobin assay. 864.7490 Section 864.7490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7490...

  13. 21 CFR 864.7425 - Carboxyhemoglobin assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Carboxyhemoglobin assay. 864.7425 Section 864.7425 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7425...

  14. 21 CFR 864.7425 - Carboxyhemoglobin assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Carboxyhemoglobin assay. 864.7425 Section 864.7425 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7425...

  15. 21 CFR 864.7490 - Sulfhemoglobin assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Sulfhemoglobin assay. 864.7490 Section 864.7490 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7490...

  16. Assay for Angiotensin-Converting Enzyme.

    ERIC Educational Resources Information Center

    Russo, Salvatore F.

    1983-01-01

    Describes a three-hour experiment designed to introduce students to chemistry of the angiotensis-converting enzyme, illustrate design of a quenched fluorescence substrate, and examine considerations necessary in designing a clinical assay. Includes background information on the biochemistry of hypertension, reagents/materials needed, procedures…

  17. Benzodiazepine Synthesis and Rapid Toxicity Assay

    ERIC Educational Resources Information Center

    Fletcher, James T.; Boriraj, Grit

    2010-01-01

    A second-year organic chemistry laboratory experiment to introduce students to general concepts of medicinal chemistry is described. Within a single three-hour time window, students experience the synthesis of a biologically active small molecule and the assaying of its biological toxicity. Benzodiazepine rings are commonly found in antidepressant…

  18. Functionalized Nanofiber Meshes Enhance Immunosorbent Assays.

    PubMed

    Hersey, Joseph S; Meller, Amit; Grinstaff, Mark W

    2015-12-01

    Three-dimensional substrates with high surface-to-volume ratios and subsequently large protein binding capacities are of interest for advanced immunosorbent assays utilizing integrated microfluidics and nanosensing elements. A library of bioactive and antifouling electrospun nanofiber substrates, which are composed of high-molecular-weight poly(oxanorbornene) derivatives, is described. Specifically, a set of copolymers are synthesized from three 7-oxanorbornene monomers to create a set of water insoluble copolymers with both biotin (bioactive) and triethylene glycol (TEG) (antifouling) functionality. Porous three-dimensional nanofiber meshes are electrospun from these copolymers with the ability to specifically bind streptavidin while minimizing the nonspecific binding of other proteins. Fluorescently labeled streptavidin is used to quantify the streptavidin binding capacity of each mesh type through confocal microscopy. A simplified enzyme-linked immunosorbent assay (ELISA) is presented to assess the protein binding capabilities and detection limits of these nanofiber meshes under both static conditions (26 h) and flow conditions (1 h) for a model target protein (i.e., mouse IgG) using a horseradish peroxidase (HRP) colorimetric assay. Bioactive and antifouling nanofiber meshes outperform traditional streptavidin-coated polystyrene plates under flow, validating their use in future advanced immunosorbent assays and their compatibility with microfluidic-based biosensors.

  19. Micropallets for cell and biological assay applications

    NASA Astrophysics Data System (ADS)

    Jensen-McMullin, Cynthia

    2007-12-01

    Interest in the subjects of microfluidics, nanotechnology and lab-on-a-chip is ever increasing. Several features of microanalysis and biological assays are desired, such as low reagent use and rapid results. These features can be achieved by developing a flexible, encoded technology capable of multiplexing. The work presented in this dissertation introduces microcarriers referred to as 'micropallets' which are encoded structures ranging in size from 25mum to several hundred microns. These small structures are fabricated using photoresist or other polymer materials. Micropallets may be used in static detection systems or for the transportation and manipulation of attached biological or chemical samples through a microfluidic system. Encoding options for micropallets are discussed. Encoding may be accomplished through the use of barcodes or other markings and may be engineered to optimally suit the application. This work presents the encoded micropallet microcarriers and the corresponding microfluidic and static systems used with micropallets. We discuss the importance of encoding towards the development of flexible, multiplexed assays and decoding strategies used or under development. Cell and antibody assays were selected and investigated to assess the utility of micropallets. We conclude from the results of this work, as well as ongoing interests, micropallets achieve the goals of improving biological techniques including cellular and other biological assays through the options of encoding and multiplexing.

  20. Advanced analysis techniques for uranium assay

    SciTech Connect

    Geist, W. H.; Ensslin, Norbert; Carrillo, L. A.; Beard, C. A.

    2001-01-01

    Uranium has a negligible passive neutron emission rate making its assay practicable only with an active interrogation method. The active interrogation uses external neutron sources to induce fission events in the uranium in order to determine the mass. This technique requires careful calibration with standards that are representative of the items to be assayed. The samples to be measured are not always well represented by the available standards which often leads to large biases. A technique of active multiplicity counting is being developed to reduce some of these assay difficulties. Active multiplicity counting uses the measured doubles and triples count rates to determine the neutron multiplication (f4) and the product of the source-sample coupling ( C ) and the 235U mass (m). Since the 35U mass always appears in the multiplicity equations as the product of Cm, the coupling needs to be determined before the mass can be known. A relationship has been developed that relates the coupling to the neutron multiplication. The relationship is based on both an analytical derivation and also on empirical observations. To determine a scaling constant present in this relationship, known standards must be used. Evaluation of experimental data revealed an improvement over the traditional calibration curve analysis method of fitting the doubles count rate to the 235Um ass. Active multiplicity assay appears to relax the requirement that the calibration standards and unknown items have the same chemical form and geometry.

  1. Calcium flux assay in Xenopus oocytes.

    PubMed

    Murphy, P M

    2001-05-01

    Many G protein-coupled receptors of interest to neuroscientists induce transient increases in [Ca(2+)](i), which can be used as a convenient measure of receptor activation in a variety of applications. This unit describes a simple calcium flux assay applied to Xenopus oocytes. PMID:18428482

  2. Three-dimensional colorimetric assay assemblies

    DOEpatents

    Charych, Deborah; Reichert, Anke

    2001-01-01

    A direct assay is described using novel three-dimensional polymeric assemblies which change from a blue to red color when exposed to an analyte, in one case a flue virus. The assemblies are typically in the form of liposomes which can be maintained in a suspension, and show great intensity in their color changes. Their method of production is also described.

  3. Homogeneous screening assay for human tankyrase.

    PubMed

    Narwal, Mohit; Fallarero, Adyary; Vuorela, Pia; Lehtiö, Lari

    2012-06-01

    Tankyrase, a member of human PARP protein superfamily, catalyzes a covalent post-translational modification of substrate proteins. This modification, poly(ADP-ribos)ylation, leads to changes in protein interactions and modifies downstream signaling events. Tankyrase 1 is a potential drug target due to its functions in telomere homeostasis and in Wnt signaling. We describe here optimization and application of an activity-based homogenous assay for tankyrase inhibitors in a high-throughput screening format. The method measures the consumption of substrate by the chemical conversion of the remaining NAD(+) into a stable fluorescent condensation product. Conditions were optimized to measure the enzymatic auto-modification of a recombinant catalytic fragment of tankyrase 1. The fluorescence assay is inexpensive, operationally easy and performs well according to the statistical analysis (Z'= 0.7). A validatory screen with a natural product library confirmed suitability of the assay for finding new tankyrase inhibitors. Flavone was the most potent (IC(50)=325 nM) hit from the natural compounds. A flavone derivative, apigenin, and isopropyl gallate showed potency on the micromolar range, but displayed over 30-fold selectivity for tankyrase over the studied isoenzymes PARP1 and PARP2. The assay is robust and will be useful for screening new tankyrase inhibitors. PMID:22357873

  4. Nondestructive assay of boxed radioactive waste

    SciTech Connect

    Gilles, W.P.; Roberts, R.J.; Jasen, W.G.

    1992-12-01

    This paper describes the problems related to the nondestructive assay (NDA) of boxed radioactive waste at the Hanford Site and how Westinghouse Hanford company (WHC) is solving the problems. The waste form and radionuclide content are described. The characteristics of the combined neutron and gamma-based measurement system are described.

  5. Analysis of Gold Ores by Fire Assay

    ERIC Educational Resources Information Center

    Blyth, Kristy M.; Phillips, David N.; van Bronswijk, Wilhelm

    2004-01-01

    Students of an Applied Chemistry degree course carried out a fire-assay exercise. The analysis showed that the technique was a worthwhile quantitative analytical technique and covered interesting theory including acid-base and redox chemistry and other concepts such as inquarting and cupelling.

  6. Surface enhanced Raman spectroscopy based nanoparticle assays for rapid, point-of-care diagnostics

    NASA Astrophysics Data System (ADS)

    Driscoll, Ashley J.

    Nucleotide and immunoassays are important tools for disease diagnostics. Many of the current laboratory-based analytical diagnostic techniques require multiple assay steps and long incubation times before results are acquired. In the development of bioassays designed for detecting the emergence and spread of diseases in point-of-care (POC) and remote settings, more rapid and portable analytical methods are necessary. Nanoparticles provide simple and reproducible synthetic methods for the preparation of substrates that can be applied in colloidal assays, providing gains in kinetics due to miniaturization and plasmonic substrates for surface enhanced spectroscopies. Specifically, surface enhanced Raman spectroscopy (SERS) is finding broad application as a signal transduction method in immunological and nucleotide assays due to the production of narrow spectral peaks from the scattering molecules and the potential for simultaneous multiple analyte detection. The application of SERS to a no-wash, magnetic capture assay for the detection of West Nile Virus Envelope and Rift Valley Fever Virus N antigens is described. The platform utilizes colloid based capture of the target antigen in solution, magnetic collection of the immunocomplexes and acquisition of SERS spectra by a handheld Raman spectrometer. The reagents for a core-shell nanoparticle, SERS based assay designed for the capture of target microRNA implicated in acute myocardial infarction are also characterized. Several new, small molecule Raman scatterers are introduced and used to analyze the enhancing properties of the synthesized gold coated-magnetic nanoparticles. Nucleotide and immunoassay platforms have shown improvements in speed and analyte capture through the miniaturization of the capture surface and particle-based capture systems can provide a route to further surface miniaturization. A reaction-diffusion model of the colloidal assay platform is presented to understand the interplay of system

  7. Development of a panel of recombinase polymerase amplification assays for detection of biothreat agents.

    PubMed

    Euler, Milena; Wang, Yongjie; Heidenreich, Doris; Patel, Pranav; Strohmeier, Oliver; Hakenberg, Sydney; Niedrig, Matthias; Hufert, Frank T; Weidmann, Manfred

    2013-04-01

    Syndromic panels for infectious disease have been suggested to be of value in point-of-care diagnostics for developing countries and for biodefense. To test the performance of isothermal recombinase polymerase amplification (RPA) assays, we developed a panel of 10 RPAs for biothreat agents. The panel included RPAs for Francisella tularensis, Yersinia pestis, Bacillus anthracis, variola virus, and reverse transcriptase RPA (RT-RPA) assays for Rift Valley fever virus, Ebola virus, Sudan virus, and Marburg virus. Their analytical sensitivities ranged from 16 to 21 molecules detected (probit analysis) for the majority of RPA and RT-RPA assays. A magnetic bead-based total nucleic acid extraction method was combined with the RPAs and tested using inactivated whole organisms spiked into plasma. The RPA showed comparable sensitivities to real-time RCR assays in these extracts. The run times of the assays at 42°C ranged from 6 to 10 min, and they showed no cross-detection of any of the target genomes of the panel nor of the human genome. The RPAs therefore seem suitable for the implementation of syndromic panels onto microfluidic platforms.

  8. Development of a Panel of Recombinase Polymerase Amplification Assays for Detection of Biothreat Agents

    PubMed Central

    Euler, Milena; Wang, Yongjie; Heidenreich, Doris; Patel, Pranav; Strohmeier, Oliver; Hakenberg, Sydney; Niedrig, Matthias; Hufert, Frank T.

    2013-01-01

    Syndromic panels for infectious disease have been suggested to be of value in point-of-care diagnostics for developing countries and for biodefense. To test the performance of isothermal recombinase polymerase amplification (RPA) assays, we developed a panel of 10 RPAs for biothreat agents. The panel included RPAs for Francisella tularensis, Yersinia pestis, Bacillus anthracis, variola virus, and reverse transcriptase RPA (RT-RPA) assays for Rift Valley fever virus, Ebola virus, Sudan virus, and Marburg virus. Their analytical sensitivities ranged from 16 to 21 molecules detected (probit analysis) for the majority of RPA and RT-RPA assays. A magnetic bead-based total nucleic acid extraction method was combined with the RPAs and tested using inactivated whole organisms spiked into plasma. The RPA showed comparable sensitivities to real-time RCR assays in these extracts. The run times of the assays at 42°C ranged from 6 to 10 min, and they showed no cross-detection of any of the target genomes of the panel nor of the human genome. The RPAs therefore seem suitable for the implementation of syndromic panels onto microfluidic platforms. PMID:23345286

  9. Production and assay of forskolin antibodies

    SciTech Connect

    Ho, L.T.; Ho, R.J.

    1986-05-01

    Forskolin (Fo), a cardiovascular active diterpene of plant origin, has been widely used as a research tool in regulation of the catalytic activity of adenylate cyclase (AC). A linear relationship of Fo binding to plasma membrane with activation of AC has been reported. The present abstract describes the production and assay of Fo antibodies (AB). 7-0-Hemisuccinyl-7-deacetyl Fo, coupled to either human serum albumin or goat IgG, was injected into goats to elicit AB to Fo haptan. AB to Fo in antiserum or an isolated IgG fraction was tested by two assay methods, a radioimmunoassay using /sup 3/H-Fo as a tracer and a colorimetric enzyme-linked immunosorbent assay (ELISA) using horse radish peroxidase-rabbit anti goat IgG as indicator. The titers for Fo antiserum were 4000-10,000. In the defined assay condition, approximately 20-25% of the added /sup 3/H-Fo was found to bind to AB. The bound radioactivity was displaced by Fo-HSA or Fo-goat IgG or free unlabelled Fo ranging from 0.5-50 pmol/tube, or 5-500 nM. The IC/sub 50/ was approximately 8-10 pmol/tube or 80-100 nM. The binding of HRP-rabbit anti goat IgG in the ELISA was inhibited by proper Fo conjugate. The development of methods for production and assay for Fo AB may be useful in the study of mechanism of activation of AC by Fo and Fo-like compound.

  10. Assays for mammalian tyrosinase: a comparative study

    SciTech Connect

    Jara, J.R.; Solano, F.; Lozano, J.A.

    1988-01-01

    This work describes a comparative study of the tyrosinase activity determined using three methods which are the most extensively employed; two radiometric assays using L-tyrosine as substrate (tyrosine hydroxylase and melanin formation activities) and one spectrophotometric assay using L-dopa (dopa oxidase activity). The three methods were simultaneously employed to measure the activities of the soluble, melanosomal, and microsomal tyrosinase isozymes from Harding-Passey mouse melanoma through their purification processes. The aim of this study was to find any correlation among the tyrosinase activities measured by the three different assays and to determine whether that correlation varied with the isozyme and its degree of purification. The results show that mammalian tyrosinase has a greater turnover number for L-dopa than for L-tyrosine. Thus, enzyme activity, expressed as mumol of substrate transformed per min, is higher in assays using L-dopa as substrate than those using L-tyrosine. Moreover, the percentage of hydroxylated L-tyrosine that is converted into melanin is low and is affected by several factors, apparently decreasing the tyrosinase activity measured by the melanin formation assay. Bearing these considerations in mind, average interassay factors are proposed. Their values are 10 to transform melanin formation into tyrosine hydroxylase activity, 100 to transform tyrosine hydroxylase into dopa oxidase activity, and 1,000 to transform melanin formation into dopa oxidase activity. Variations in these values due to the presence in the tyrosinase preparations of either inhibitors or regulatory factors in melanogenesis independent of tyrosinase are also discussed.

  11. Comet assay: rapid processing of multiple samples.

    PubMed

    McNamee, J P; McLean, J R; Ferrarotto, C L; Bellier, P V

    2000-03-01

    The present study describes modifications to the basic comet protocol that increase productivity and efficiency without sacrificing assay reliability. A simple technique is described for rapidly preparing up to 96 comet assay samples simultaneously. The sample preparation technique allows thin layers of agarose-embedded cells to be prepared in multiple wells attached to a flexible film of Gelbond, which improves the ease of manipulating and processing samples. To evaluate the effect of these modifications on assay sensitivity, dose-response curves are presented for DNA damage induced by exposure of TK6 cells to low concentrations of hydrogen peroxide (0-10 microM) and for exposure of human lymphocytes to X-irradiation (0-100 cGy). The limit of detection of DNA damage induced by hydrogen peroxide in TK6 cells was observed to be 1 uM for all parameters (tail ratio, tail moment, tail length and comet length) while the limit of detection of DNA damage in human lymphocytes was 10 cGy for tail and comet length parameters, but 50 cGy for tail ratio and tail moment parameters. These results are similar to those previously reported using the conventional alkaline comet assay. The application of SYBR Gold for detection of DNA damage was compared to that of propidium iodide. Measurements of matching samples for tail length and comet length were similar using both stains. However, comets stained with SYBR Gold persisted longer and were much brighter than those obtained with propidium iodide. SYBR Gold was found to be ideal for measuring tail length and comet length but, under present assay conditions, impractical for measuring tail ratio or tail moment due to saturation of staining in the head region of the comets. PMID:10751727

  12. Non-separation assay for glycohemoglobin.

    PubMed

    Blincko, S; Edwards, R

    1998-06-01

    The determination of glycohemoglobin [HbA1c, HbA1, or total glycohemoglobin (GHb)] has become an established procedure in the management of diabetes mellitus. Here, we describe the development of a simple, fluorescence, non-separation assay for the percentage of GHb (%GHb). The fluorescence of an eosin-boronic acid derivative when it was mixed with hemolysates of unwashed erythrocytes was quenched in proportion to the percentage of glycohemoglobin. Measurement of the fluorescence intensity gave an estimate of GHb in the sample, and measurement of light absorbance gave an estimate of total hemoglobin. A combination of the two measurements gave the assay response. Comparison with HPLC (Menarini-Arkray HA-8140 fully automated analyzer) for the percentage of HbA1 (%HbA1) gave %GHb(NETRIA) = 1.1(SD +/-0.03)%HbA1 +0.6(SD +/-0.3), S(y/x) = 0.821, r = 0.972, n = 80; comparison for HbA1c gave %GHb(NETRIA) = 1.3(SD +/-0.04)%HbA1c + 1.8(SD +/-0.3), S(y/x) = 0.813, r = 0.973, n = 80. Precision, estimated as the percentage of the CV of the %GHb assay results, was <2% (intraassay, range 5-22% GHb) and <4.2% (interassay, range 4-16% GHb). Dilution of a high-percentage GHb sample lysate showed that the assay was linear, and addition of glucose (60 mmol/L), bilirubin (250 micromol/L), and triglycerides (14 mmol/L) to low, medium, and high %GHb samples showed no clinical interference in assay results. PMID:9625057

  13. Detection of two different influenza A viruses using a nitrocellulose membrane and a magnetic biosensor.

    PubMed

    Hong, Hyo-Bong; Krause, Hans-Joachim; Song, Ki-Bong; Choi, Chel-Jong; Chung, Myung-Ae; Son, Sung-Won; Offenhäusser, Andreas

    2011-02-28

    Here we describe a new analytical method for the detection of two influenza A viruses by nitrocellulose membrane and magnetic sensors that employ a special frequency mixing technique. The combination of the nitrocellulose membrane and magnetic bead detection permits a rapid assay procedure and excludes two steps (the development of color and the stop reaction) required for usual immunochemical detection methods such as ELISA. Quantitative virus detection was performed using magnetic beads conjugated with secondary antibody. The results were compared with conventional assay methods and with a dot-blot assay with fluorescence compound (FITC). Under optimum conditions, our new assay procedure is capable of detecting picograms of virus per well. This new method combining the nitrocellulose membrane and magnetic bead detection reduces analytical time and allows stable and repeatable analyses of samples in point-of-care applications.

  14. Variola Virus-Specific Diagnostic Assays: Characterization, Sensitivity, and Specificity

    PubMed Central

    Kondas, Ashley V.; Olson, Victoria A.; Li, Yu; Abel, Jason; Laker, Miriam; Rose, Laura; Wilkins, Kimberly; Turner, Jonathan; Kline, Richard

    2015-01-01

    A public health response relies upon rapid and reliable confirmation of disease by diagnostic assays. Here, we detail the design and validation of two variola virus-specific real-time PCR assays, since previous assays cross-reacted with newly identified cowpox viruses. The assay specificity must continually be reassessed as other closely related viruses are identified. PMID:25673790

  15. Variola virus-specific diagnostic assays: characterization, sensitivity, and specificity.

    PubMed

    Kondas, Ashley V; Olson, Victoria A; Li, Yu; Abel, Jason; Laker, Miriam; Rose, Laura; Wilkins, Kimberly; Turner, Jonathan; Kline, Richard; Damon, Inger K

    2015-04-01

    A public health response relies upon rapid and reliable confirmation of disease by diagnostic assays. Here, we detail the design and validation of two variola virus-specific real-time PCR assays, since previous assays cross-reacted with newly identified cowpox viruses. The assay specificity must continually be reassessed as other closely related viruses are identified.

  16. Ferroelectricity in spiral magnets.

    PubMed

    Mostovoy, Maxim

    2006-02-17

    It was recently observed that the ferroelectrics showing the strongest sensitivity to an applied magnetic field are spiral magnets. We present a phenomenological theory of inhomogeneous ferroelectric magnets, which describes their thermodynamics and magnetic field behavior, e.g., dielectric susceptibility anomalies at magnetic transitions and sudden flops of electric polarization in an applied magnetic field. We show that electric polarization can also be induced at domain walls and that magnetic vortices carry electric charge. PMID:16606047

  17. A versatile electrowetting-based digital microfluidic platform for quantitative homogeneous and heterogeneous bio-assays

    NASA Astrophysics Data System (ADS)

    Vergauwe, Nicolas; Witters, Daan; Ceyssens, Frederik; Vermeir, Steven; Verbruggen, Bert; Puers, Robert; Lammertyn, Jeroen

    2011-05-01

    Electrowetting-on-dielectric (EWOD) lab-on-a-chip systems have already proven their potential within a broad range of bio-assays. Nevertheless, research on the analytical performance of those systems is limited, yet crucial for a further breakthrough in the diagnostic field. Therefore, this paper presents the intrinsic possibilities of an EWOD lab-on-a-chip as a versatile platform for homogeneous and heterogeneous bio-assays with high analytical performance. Both droplet dispensing and splitting cause variations in droplet size, thereby directly influencing the assay's performance. The extent to which they influence the performance is assessed by a theoretical sensitivity analysis, which allows the definition of a basic framework for the reduction of droplet size variability. Taking advantage of the optimized droplet manipulations, both homogeneous and heterogeneous bio-assays are implemented in the EWOD lab-on-a-chip to demonstrate the analytical capabilities and versatility of the device. A fully on-chip enzymatic assay is realized with high analytical performance. It demonstrates the promising capabilities of an EWOD lab-on-a-chip in food-related and medical applications, such as nutritional and blood analyses. Further, a magnetic bio-assay for IgE detection using superparamagnetic nanoparticles is presented whereby the nanoparticles are used as solid carriers during the bio-assay. Crucial elements are the precise manipulation of the superparamagnetic nanoparticles with respect to dispensing and separation. Although the principle of using nano-carriers is demonstrated for protein detection, it can be easily extended to a broader range of bio-related applications like DNA sensing. In heterogeneous bio-assays the chip surface is actively involved during the execution of the bio-assay. Through immobilization of specific biological compounds like DNA, proteins and cells a reactive chip surface is realized, which enhances the bio-assay performance. To demonstrate

  18. Fast Magnetic Field-Enhanced Linear Colloidal Agglutination Immunoassay.

    PubMed

    Daynès, Aurélien; Temurok, Nevzat; Gineys, Jean-Philippe; Cauet, Gilles; Nerin, Philippe; Baudry, Jean; Bibette, Jérôme

    2015-08-01

    We present the principle of a fast magnetic field enhanced colloidal agglutination assay, which is based on the acceleration of the recognition rate between ligands and receptors induced by magnetic forces. By applying a homogeneous magnetic field of 20 mT for only 7 s, we detect CRP (C-reactive protein) in human serum at a concentration as low as 1 pM for a total cycle time of about 1 min in a prototype analyzer. Such a short measurement time does not impair the performances of the assay when compared to longer experiments. The concentration range dynamic is shown to cover 3 orders of magnitude. An analytical model of agglutination is also successfully fitting our data obtained with a short magnetic pulse.

  19. Storage and Assay of Tritium in STAR

    SciTech Connect

    Longhurst, Glen R.; Anderl, Robert A.; Pawelko, Robert J.; Stoots, Carl J.

    2005-07-15

    The Safety and Tritium Applied Research (STAR) facility at the Idaho National Engineering and Environmental Laboratory (INEEL) is currently being commissioned to investigate tritium-related safety questions for fusion and other technologies. The tritium inventory for the STAR facility will be maintained below 1.5 g to avoid the need for STAR to be classified as a Category 3 nuclear facility. A key capability in successful operation of the STAR facility is the ability to receive, inventory, and dispense tritium to the various experiments underway there. The system central to that function is the Tritium Storage and Assay System (SAS).The SAS has four major functions: (1) receiving and holding tritium, (2) assaying, (3) dispensing, and (4) purifying hydrogen isotopes from non-hydrogen species.This paper describes the design and operation of the STAR SAS and the procedures used for tritium accountancy in the STAR facility.

  20. Bio-assays for microchemical environmental contaminants

    PubMed Central

    Warner, Richard E.

    1967-01-01

    A solution of the problem of environmental contamination must be based on accurate measurement of the extent of the contamination and of the resulting hazards. This paper reviews the methods for the estimation of microchemical contaminants in water with the aid of living organisms. The methods are grouped according to the nature of the response of the organism to the contaminant—namely, acute response (usually death), behavioural change, physiological change, biochemical and histochemical change, ecological change, embryological and regenerational change, growth change, histological change and perception by man or aquatic organisms. Finally, the following problems are discussed: selection of appropriate tests and standardization, the dangers of sequential concentration and the need for multi-parametric assays (assays involving several responses of a single organism, or responses of several organisms) for complete characterization of the effects of a contaminant on the environment. ImagesFIG. 2FIG. 3FIG. 4FIG. 5FIG. 6 PMID:5299747

  1. Detailed assays conducted on Vietnamese crude oils

    SciTech Connect

    Du, P.Q. )

    1990-07-16

    More oil property data, in the form of recent crude oil assays, have been made available for two Vietnamese crude oils, Bach Ho (White Tiger) and Dai Hung (Big Bear). Crude oil data presented earlier gave limited properties of the crudes,which are from the Miocene formations. Further analyses have been conducted on Bach Ho crude from the Oligocene formations. Production from Oligocene is far more representative of the oils produced from the Bach Ho field and marketed worldwide. Currently, Bach Ho is the only producing field. Dai Hung is expected to be in production during the next few years. Bach Ho is currently producing at the rate of 20,000 b/d. That figure is projected to grow to 100,000 b/d by 1992 and to 120,000 b/d by 1995. Detailed assays of both crude oils are presented.

  2. Expert system technology for nondestructive waste assay

    SciTech Connect

    Becker, G.K.; Determan, J.C.

    1998-07-01

    Nondestructive assay waste characterization data generated for use in the National TRU Program must be of known and demonstrable quality. Each measurement is required to receive an independent technical review by a qualified expert. An expert system prototype has been developed to automate waste NDA data review of a passive/active neutron drum counter system. The expert system is designed to yield a confidence rating regarding measurement validity. Expert system rules are derived from data in a process involving data clustering, fuzzy logic, and genetic algorithms. Expert system performance is assessed against confidence assignments elicited from waste NDA domain experts. Performance levels varied for the active, passive shielded, and passive system assay modes of the drum counter system, ranging from 78% to 94% correct classifications.

  3. A specific endpoint assay for ubiquitin.

    PubMed Central

    Rose, I A; Warms, J V

    1987-01-01

    Simple endpoint assays for free ubiquitin (Ub) and for the Ub-activating enzyme are described. The method for measuring Ub makes use of the reaction of iodoacetamide-treated Ub-activating enzyme (E): [3H]ATP + Ub + E----E X [3H]AMP-Ub + PPi and PPi----2Pi (in the presence of pyrophosphatase). The Ub is then measured by determining the acid-insoluble radioactivity. The reaction is accompanied by a slow enzyme-catalyzed hydrolysis of the complex to AMP plus Ub. The presence of ubiquitin-activating enzyme in excess of Ub by approximately equal to 0.1 microM assures that the steady state will be close to the endpoint for total Ub. A preparation of the activating enzyme from human erythrocytes that does not depend on affinity chromatography is described. Several applications of the assay are presented. PMID:3031643

  4. Immunonephelometric and immunoturbidimetric assays for proteins.

    PubMed

    Whicher, J T; Price, C P; Spencer, K

    1983-01-01

    Immunonephelometric and immunoturbidimetric techniques for the measurement of proteins have developed and expanded rapidly in recent years and are fast replacing the time-honoured gel precipitation techniques. The reasons for this are primarily an increased awareness of the value of specific protein measurements with the impetus of improved chemical reagents and advances in instrument technology. This review discusses the background to such free fluid phase immunochemical measurement systems and the developments which have occurred in this field. The following topics are reviewed. (1) The theoretical background of light scattering theory as applied to the measurement of antibody-antigen complexes. (2) The nature and kinetics of the antibody-antigen reaction in fluid media and the effect of enhancing polymers. (3) The sample and antibody requirements for nephelometric and turbidimetric assay. (4) Instrumental systems for nephelometry. (5) Instrumental systems for turbidimetry. (6) Methods of establishing, assessing and monitoring nephelometric assays for specific proteins in the laboratory.

  5. Quantum Dot-Based Cell Motility Assay

    SciTech Connect

    Gu, Weiwei; Pellegrino, Teresa; Parak Wolfgang J; Boudreau,Rosanne; Le Gros, Mark A.; Gerion, Daniele; Alivisatos, A. Paul; Larabell, Carolyn A.

    2005-06-06

    Because of their favorable physical and photochemical properties, colloidal CdSe/ZnS-semiconductor nanocrystals (commonly known as quantum dots) have enormous potential for use in biological imaging. In this report, we present an assay that uses quantum dots as markers to quantify cell motility. Cells that are seeded onto a homogeneous layer of quantum dots engulf and absorb the nanocrystals and, as a consequence, leave behind a fluorescence-free trail. By subsequently determining the ratio of cell area to fluorescence-free track area, we show that it is possible to differentiate between invasive and noninvasive cancer cells. Because this assay uses simple fluorescence detection, requires no significant data processing, and can be used in live-cell studies, it has the potential to be a powerful new tool for discriminating between invasive and noninvasive cancer cell lines or for studying cell signaling events involved in migration.

  6. Delayed gamma technique for fissile material assay

    SciTech Connect

    Mozin, Vladimir; Tobin, Stephen; Vujie, Jasmina; Hunt, Alan

    2010-01-01

    Research sponsored by the Next Generation Safeguards Initiative are investigating several non-destructive assay techniques for the quantification of fissile plutonium mass in spent nuclear fuel assemblies. AppHcation of the delayed gamma signatures is investigated in this context. The objective of the research is to assess whether the delayed gamma assay instrument can provide sufficient sensitivity, isotope specificity and accuracy as required in nuclear material safeguards. This effort includes theoretical and experimental components for the optimal combination of interrogation parameters. A new modeling algorithm offering a high level of detail was developed specifically for this purpose and was validated in series of benchmark experiments. Preliminary modeling of the delayed gamma response from spent fuel assemblies was accomplished offering a future direction for the design process.

  7. Taste Preference Assay for Adult Drosophila.

    PubMed

    Bantel, Andrew P; Tessier, Charles R

    2016-01-01

    Olfactory and gustatory perception of the environment is vital for animal survival. The most obvious application of these chemosenses is to be able to distinguish good food sources from potentially dangerous food sources. Gustation requires physical contact with a chemical compound which is able to signal through taste receptors that are expressed on the surface of neurons. In insects, these gustatory neurons can be located across the animal's body allowing taste to play an important role in many different behaviors. Insects typically prefer compounds containing sugars, while compounds that are considered bitter tasting are avoided. Given the basic biological importance of taste, there is intense interest in understanding the molecular mechanisms underlying this sensory modality. We describe an adult Drosophila taste assay which reflects the preference of the animals for a given tastant compound. This assay may be applied to animals of any genetic background to examine the taste preference for a desired soluble compound. PMID:27684591

  8. Depleted uranium waste assay at AWE

    SciTech Connect

    Miller, T.J.

    2007-07-01

    The Atomic Weapons Establishment (AWE) at Aldermaston has recently conducted a Best Practical Means (BPM) study, for solid Depleted Uranium (DU) waste assay, in order to satisfy key stakeholders that AWE is applying best practice. This study has identified portable passive High Resolution Gamma Spectrometry (HRGS), combined with an analytical software package called Spectral Nondestructive Assay Platform (SNAP), as the preferred option with the best balance between performance and costs. HRGS/SNAP performance has been assessed by monitoring 200 l DU waste drum standards and also heterogeneous, high density drums from DU firing trials. Accuracy was usually within 30 % with Detection Limits (DL) in the region of 10 g DU for short count times. Monte Carlo N-Particle (MCNP) calculations have been used to confirm the shape of the calibration curve generated by the SNAP software procured from Eberline Services Inc. (authors)

  9. Neutron Assay System for Confinement Vessel Disposition

    SciTech Connect

    Frame, Katherine C.; Bourne, Mark M.; Crooks, William J.; Evans, Louise; Mayo, Douglas R.; Miko, David K.; Salazar, William R.; Stange, Sy; Valdez, Jose I.; Vigil, Georgiana M.

    2012-07-13

    Los Alamos National Laboratory has a number of spherical confinement vessels (CVs) remaining from tests involving nuclear materials. These vessels have an inner diameter of 6 feet with 1-inch thick steel walls. The goal of the Confinement Vessel Disposition (CVD) project is to remove debris and reduce contamination inside the CVs. The Confinement Vessel Assay System (CVAS) was developed to measure the amount of special nuclear material (SNM) in CVs before and after cleanout. Prior to cleanout, the system will be used to perform a verification measurement of each vessel. After cleanout, the system will be used to perform safeguards-quality assays of {le}100-g {sup 239}Pu equivalent in a vessel for safeguards termination. The CVAS has been tested and calibrated in preparation for verification and safeguards measurements.

  10. System and method for assaying a radionuclide

    SciTech Connect

    Cadieux, James R; King, III, George S; Fugate, Glenn A

    2014-12-23

    A system for assaying a radionuclide includes a liquid scintillation detector, an analyzer connected to the liquid scintillation detector, and a delay circuit connected to the analyzer. A gamma detector and a multi-channel analyzer are connected to the delay circuit and the gamma detector. The multi-channel analyzer produces a signal reflective of the radionuclide in the sample. A method for assaying a radionuclide includes selecting a sample, detecting alpha or beta emissions from the sample with a liquid scintillation detector, producing a first signal reflective of the alpha or beta emissions, and delaying the first signal a predetermined time. The method further includes detecting gamma emissions from the sample, producing a second signal reflective of the gamma emissions, and combining the delayed first signal with the second signal to produce a third signal reflective of the radionuclide.

  11. DREAM Assay for Studying Microbial Electron Transfer.

    PubMed

    Vishwanathan, A S; Devkota, Ranjan; Siva Sankara Sai, S; Rao, Govind

    2015-12-01

    Methylene blue undergoes reduction with an accompanying colour change reaction, from blue to colourless, enabling its use as a metric for estimating reducing power. A dye reduction-based electron-transfer activity monitoring (DREAM) assay is demonstrated as a tool to study and understand the process of microbes sourcing electrons from organic substrates and transferring them to an electron acceptor. The rate at which electrons can be transferred to the thermodynamically most feasible electron acceptor directly depends on the activity of microbes. Nature of available substrate determines the quantum of electrons available. Dissolved oxygen intercepts electrons from the microbes before they can be taken up by the dye. Sodium sulfite can be used to offset the detrimental effects of the presence of dissolved oxygen. This easy-to-perform assay has been demonstrated as a proof-of-concept having potential to be extended to other practical applications. PMID:26386586

  12. Assay and Inhibition of Diacylglycerol Lipase Activity

    PubMed Central

    Johnston, Meghan; Bhatt, Shachi R.; Sikka, Surina; Mercier, Richard W.; West, Jay M.; Makriyannis, Alexandros; Gatley, S. John; Duclos, Richard I.

    2012-01-01

    A series of N-formyl-α-amino acid esters of β-lactone derivatives structurally related to tetrahydrolipstatin (THL) and O-3841 were synthesized that inhibit human and murine diacylglycerol lipase (DAGL) activities. New ether lipid reporter compounds were developed for an in vitro assay to efficiently screen inhibitors of 1,2-diacyl-sn-glycerol hydrolysis and related lipase activities using fluorescence resonance energy transfer (FRET). A standardized thin layer chromatography (TLC) radioassay of diacylglycerol lipase activity utilizing the labeled endogenous substrate [1″-14C]1-stearoyl-2-arachidonoyl-sn-glycerol with phosphorimaging detection was used to quantify inhibition by following formation of the initial product [1″-14C]2-arachidonoylglycerol and further hydrolysis under the assay conditions to [1-14C]arachidonic acid. PMID:22738638

  13. [Thrombin generation assays and their clinical application].

    PubMed

    Kern, Anita; Várnai, Katalin; Vásárhelyi, Barna

    2014-06-01

    Thrombin is a key enzyme of the coagulation system, having both pro- and anticoagulant functions. Thus, the generation of thrombin is one of the most important steps in coagulation. Global haemostasis assay, the so-called thrombin generation test is appropriate for its assessment. Since thrombin generation is sensible for both pro- and anticoagulant processes it can be applied for the general characterisation of the risk of thrombosis and bleeding, too. Clinical studies confirmed augmented thrombin generation in patients with high risk of venous or arterial thrombosis. Anticoagulant therapy (also novel oral anticoagulant treatment) can be monitored by thrombin generation. In case of haemophilia thrombin generation assays reflect bleeding severity. It is applicable for monitoring of both conventional haemophilia treatment and inhibitor-bypassing therapy, which is needed when inhibitors develop in patients. Standardization of thrombin generation methods and determination of cut off values are required before its application in clinical practice.

  14. System and method for assaying radiation

    SciTech Connect

    DiPrete, David P; Whiteside, Tad; Pak, Donald J; DiPrete, Cecilia C

    2013-11-12

    A system for assaying radiation includes a sample holder configured to hold a liquid scintillation solution. A photomultiplier receives light from the liquid scintillation solution and generates a signal reflective of the light. A control circuit biases the photomultiplier and receives the signal from the photomultiplier reflective of the light. A light impermeable casing surrounds the sample holder, photomultiplier, and control circuit. A method for assaying radiation includes placing a sample in a liquid scintillation solution, placing the liquid scintillation solution in a sample holder, and placing the sample holder inside a light impermeable casing. The method further includes positioning a photomultiplier inside the light impermeable casing and supplying power to a control circuit inside the light impermeable casing.

  15. DREAM Assay for Studying Microbial Electron Transfer.

    PubMed

    Vishwanathan, A S; Devkota, Ranjan; Siva Sankara Sai, S; Rao, Govind

    2015-12-01

    Methylene blue undergoes reduction with an accompanying colour change reaction, from blue to colourless, enabling its use as a metric for estimating reducing power. A dye reduction-based electron-transfer activity monitoring (DREAM) assay is demonstrated as a tool to study and understand the process of microbes sourcing electrons from organic substrates and transferring them to an electron acceptor. The rate at which electrons can be transferred to the thermodynamically most feasible electron acceptor directly depends on the activity of microbes. Nature of available substrate determines the quantum of electrons available. Dissolved oxygen intercepts electrons from the microbes before they can be taken up by the dye. Sodium sulfite can be used to offset the detrimental effects of the presence of dissolved oxygen. This easy-to-perform assay has been demonstrated as a proof-of-concept having potential to be extended to other practical applications.

  16. Transient expression assays in tobacco protoplasts.

    PubMed

    Vanden Bossche, Robin; Demedts, Brecht; Vanderhaeghen, Rudy; Goossens, Alain

    2013-01-01

    The sequence information generated through genome and transcriptome analysis from plant tissues has reached unprecedented sizes. Sequence homology-based annotations may provide hints for the possible function and roles of particular plant genes, but the functional annotation remains nonexistent or incomplete for many of them. To discover gene functions, transient expression assays are a valuable tool because they can be done more rapidly and at a higher scale than generating stably transformed tissues. Here, we describe a transient expression assay in protoplasts derived from suspension cells of tobacco (Nicotiana tabacum) for the study of the transactivation capacities of transcription factors. To enhance throughput and reproducibility, this method can be automated, allowing medium-throughput screening of interactions between large compendia of potential transcription factors and gene promoters.

  17. Developments in plutonium waste assay at AWE.

    PubMed

    Miller, T J

    2009-06-01

    In 2002 a paper was presented at the 43rd Annual Meeting of the Institute of Nuclear Materials Management (INMM) on the assay of low level plutonium (Pu) in soft drummed waste (Miller 2002 INMM Ann. Meeting (Orlando, FL, 23-27 July 2002)). The technique described enabled the Atomic Weapons Establishment (AWE), at Aldermaston in the UK, to meet the stringent Low Level Waste Repository at Drigg (LLWRD) conditions for acceptance for the first time. However, it was initially applied to only low density waste streams because it relied on measuring the relatively low energy (60 keV) photon yield from Am-241 during growth. This paper reviews the results achieved when using the technique to assay over 10,000 waste packages and presents the case for extending the range of application to denser waste streams.

  18. Label-free functional selectivity assays.

    PubMed

    Ferrie, Ann M; Goral, Vasiliy; Wang, Chaoming; Fang, Ye

    2015-01-01

    G protein-coupled receptors (GPCRs) represent the largest class of drug targets. Ligand-directed functional selectivity or biased agonism opens new possibility for discovering GPCR drugs with better efficacy and safety profiles. However, quantification of ligand bias is challenging. Herein, we present five different label-free dynamic mass redistribution (DMR) approaches to assess ligand bias acting at the β2-adrenergic receptor (β2AR). Multiparametric analysis of the DMR agonist profiles reveals divergent pharmacology of a panel of β2AR agonists. DMR profiling using catechol as a conformational probe detects the presence of multiple conformations of the β2AR. DMR assays under microfluidics, together with chemical biology tools, discover ligand-directed desensitization of the receptor. DMR antagonist reverse assays manifest biased antagonism. DMR profiling using distinct probe-modulated cells detects the biased agonism in the context of self-referenced pharmacological activity map. PMID:25563188

  19. Assay of brines for common radiolysis products

    SciTech Connect

    MacDougall, C.S.

    1981-01-01

    Brines are assayed for four common products of radiolytic reaction. Free chlorine is determined spectrophotometrically after reaction with o-tolidine. The test is specific for chlorine, and quantities of chlorine from 0.1 to 6 ..mu..g in the test aliquot are determined with a precision of about +- 5%. Hydrogen peroxide is reacted with xylenol orange and determined spectrophotometrically with a precision of +- 5% on 2-..mu..g quantities of peroxide. A spectrophotometric method using thiocyanate is employed in the chlorate assay. After subtracting the bias caused by any H/sub 2/O/sub 2/ or Cl/sub 2/, 1-..mu..g quantities of chlorate can be determined with a precision of +- 10%. Perchlorate ion quantities of 1 ppM can be determined directly in brines by ion chromatography with a precision of about +- 15%.

  20. Biomimetic magnetic silk scaffolds.

    PubMed

    Samal, Sangram K; Dash, Mamoni; Shelyakova, Tatiana; Declercq, Heidi A; Uhlarz, Marc; Bañobre-López, Manuel; Dubruel, Peter; Cornelissen, Maria; Herrmannsdörfer, Thomas; Rivas, Jose; Padeletti, Giuseppina; De Smedt, Stefaan; Braeckmans, Kevin; Kaplan, David L; Dediu, V Alek

    2015-03-25

    Magnetic silk fibroin protein (SFP) scaffolds integrating magnetic materials and featuring magnetic gradients were prepared for potential utility in magnetic-field assisted tissue engineering. Magnetic nanoparticles (MNPs) were introduced into SFP scaffolds via dip-coating methods, resulting in magnetic SFP scaffolds with different strengths of magnetization. Magnetic SFP scaffolds showed excellent hyperthermia properties achieving temperature increases up to 8 °C in about 100 s. The scaffolds were not toxic to osteogenic cells and improved cell adhesion and proliferation. These findings suggest that tailored magnetized silk-based biomaterials can be engineered with interesting features for biomaterials and tissue-engineering applications.

  1. Electrically Tunable Magnetism in Magnetic Topological Insulators.

    PubMed

    Wang, Jing; Lian, Biao; Zhang, Shou-Cheng

    2015-07-17

    The external controllability of the magnetic properties in topological insulators would be important both for fundamental and practical interests. Here we predict the electric-field control of ferromagnetism in a thin film of insulating magnetic topological insulators. The decrease of band inversion by the application of electric fields results in a reduction of magnetic susceptibility, and hence in the modification of magnetism. Remarkably, the electric field could even induce the magnetic quantum phase transition from ferromagnetism to paramagnetism. We further propose a transistor device in which the dissipationless charge transport of chiral edge states is controlled by an electric field. In particular, the field-controlled ferromagnetism in a magnetic topological insulator can be used for voltage based writing of magnetic random access memories in magnetic tunnel junctions. The simultaneous electrical control of magnetic order and chiral edge transport in such devices may lead to electronic and spintronic applications for topological insulators.

  2. Magnetic Storms

    NASA Technical Reports Server (NTRS)

    Tsurutani, Bruce T.; Gonzalez, Walter D.

    1998-01-01

    One of the oldest mysteries in geomagnetism is the linkage between solar and geomagnetic activity. The 11-year cycles of both the numbers of sunspots and Earth geomagnetic storms were first noted by Sabine. A few years later, speculation on a causal relationship between flares and storms arose when Carrington reported that a large magnetic storm followed the great September 1859 solar flare. However, it was not until this century that a well-accepted statistical survey on large solar flares and geomagnetic storms was performed, and a significant correlation between flares and geomagnetic storms was noted. Although the two phenomena, one on the Sun and the other on the Earth, were statistically correlated, the exact physical linkage was still an unknown at this time. Various hypotheses were proposed, but it was not until interplanetary spacecraft measurements were available that a high-speed plasma stream rich in helium was associated with an intense solar flare. The velocity of the solar wind increased just prior to and during the helium passage, identifying the solar ejecta for the first time. Space plasma measurements and Skylab's coronagraph images of coronal mass elections (CMES) from the Sun firmly established the plasma link between the Sun and the Earth. One phenomenon associated with magnetic storms is brilliant "blood" red auroras, as shown.

  3. Superconducting magnet

    DOEpatents

    Satti, John A.

    1980-01-01

    A superconducting magnet designed to produce magnetic flux densities of the order of 4 to 5 Webers per square meter is constructed by first forming a cable of a plurality of matrixed superconductor wires with each wire of the plurality insulated from each other one. The cable is shaped into a rectangular cross-section and is wound with tape in an open spiral to create cooling channels. Coils are wound in a calculated pattern in saddle shapes to produce desired fields, such as dipoles, quadrupoles, and the like. Wedges are inserted between adjacent cables as needed to maintain substantially radial placement of the long dimensions of cross sections of the cables. After winding, individual strands in each of the cables are brought out to terminals and are interconnected to place all of the strands in series and to maximize the propagation of a quench by alternating conduction from an inner layer to an outer layer and from top half to bottom half as often as possible. Individual layers are separated from others by spiraled aluminum spacers to facilitate cooling. The wound coil is wrapped with an epoxy tape that is cured by heat and then machined to an interference fit with an outer aluminum pipe which is then affixed securely to the assembled coil by heating it to make a shrink fit. In an alternate embodiment, one wire of the cable is made of copper or the like to be heated externally to propagate a quench.

  4. Quantifiable Lateral Flow Assay Test Strips

    NASA Technical Reports Server (NTRS)

    2003-01-01

    As easy to read as a home pregnancy test, three Quantifiable Lateral Flow Assay (QLFA) strips used to test water for E. coli show different results. The brightly glowing control line on the far right of each strip indicates that all three tests ran successfully. But the glowing test line on the middle left and bottom strips reveal their samples were contaminated with E. coli bacteria at two different concentrations. The color intensity correlates with concentration of contamination.

  5. Polymeric assay film for direct colorimetric detection

    DOEpatents

    Charych, Deborah; Nagy, Jon; Spevak, Wayne

    2002-01-01

    A lipid bilayer with affinity to an analyte, which directly signals binding by a changes in the light absorption spectra. This novel assay means and method has special applications in the drug development and medical testing fields. Using a spectrometer, the system is easily automated, and a multiple well embodiment allows inexpensive screening and sequential testing. This invention also has applications in industry for feedstock and effluent monitoring.

  6. Polymeric assay film for direct colorimetric detection

    DOEpatents

    Charych, Deborah; Nagy, Jon; Spevak, Wayne

    1999-01-01

    A lipid bilayer with affinity to an analyte, which directly signals binding by a changes in the light absorption spectra. This novel assay means and method has special applications in the drug development and medical testing fields. Using a spectrometer, the system is easily automated, and a multiple well embodiment allows inexpensive screening and sequential testing. This invention also has applications in industry for feedstock and effluent monitoring.

  7. Methods and devices for protein assays

    DOEpatents

    Chhabra, Swapnil; Cintron, Jose M.; Shediac, Renee

    2009-11-03

    Methods and devices for protein assays based on Edman degradation in microfluidic channels are disclosed herein. As disclosed, the cleaved amino acid residues may be immobilized in an array format and identified by detectable labels, such as antibodies, which specifically bind given amino acid residues. Alternatively, the antibodies are immobilized in an array format and the cleaved amino acids are labeled identified by being bound by the antibodies in the array.

  8. Development of Magnetic Refrigerator

    NASA Astrophysics Data System (ADS)

    Ogiwara, Hiroyasu; Nakagome, Hideki; Kuriyama, Tohru

    A series of R & D of magnetic refrigerators has been done in order to realize an advanced type cryocooler for superconducting magnets of maglev trains and MRI medical system. As a result of efforts on both the magnetic refrigerator and superconducting magnets, a parasitic type magnetic refrigeration system was proposed.

  9. Permanent magnet assembly

    DOEpatents

    Chell, Jeremy; Zimm, Carl B.

    2006-12-12

    A permanent magnet assembly is disclosed that is adapted to provide a magnetic field across an arc-shaped gap. Such a permanent magnet assembly can be used, for example, to provide a time-varying magnetic field to an annular region for use in a magnetic refrigerator.

  10. The magnetization process: Hysteresis

    NASA Technical Reports Server (NTRS)

    Balsamel, Richard

    1990-01-01

    The magnetization process, hysteresis (the difference in the path of magnetization for an increasing and decreasing magnetic field), hysteresis loops, and hard magnetic materials are discussed. The fabrication of classroom projects for demonstrating hysteresis and the hysteresis of common magnetic materials is described in detail.

  11. Magnet innovations for linacs

    SciTech Connect

    Halbach, K.

    1986-06-01

    It is possible to produce large magnetic fields at the aperture of permanent magnet quadrupoles, even when the magnetic aperture is very small. That, combined with their compactness, makes permanent magnet quadrupoles very powerful components of small aperture linacs. Results will be presented about past and present work on both fixed and variable strength permanent magnets suitable for use in and around linacs.

  12. Magnetic field sensor

    NASA Astrophysics Data System (ADS)

    Silva, Nicolas

    2012-09-01

    Earlier papers1-3 in this journal have described experiments on measuring the magnetic fields of current-carrying wires and permanent magnets using magnetic field probes of various kinds. This paper explains how to use an iPad and the free app MagnetMeter-3D Vector Magnetometer and Accelerometer4 (compass HD) to measure the magnetic fields.

  13. Lymphocyte chromosomal aberration assay in radiation biodosimetry

    PubMed Central

    Agrawala, Paban K.; Adhikari, J. S.; Chaudhury, N. K.

    2010-01-01

    Exposure to ionizing radiations, whether medical, occupational or accidental, leads to deleterious biological consequences like mortality or carcinogenesis. It is considered that no dose of ionizing radiation exposure is safe. However, once the accurate absorbed dose is estimated, one can be given appropriate medical care and the severe consequences can be minimized. Though several accurate physical dose estimation modalities exist, it is essential to estimate the absorbed dose in biological system taking into account the individual variation in radiation response, so as to plan suitable medical care. Over the last several decades, lots of efforts have been taken to design a rapid and easy biological dosimeter requiring minimum invasive procedures. The metaphase chromosomal aberration assay in human lymphocytes, though is labor intensive and requires skilled individuals, still remains the gold standard for radiation biodosimetry. The current review aims at discussing the human lymphocyte metaphase chromosomal aberration assay and recent developments involving the application of molecular cytogenetic approaches and other technological advancements to make the assay more authentic and simple to use even in the events of mass radiation casualties. PMID:21829315

  14. Assessment of plaque assay methods for alphaviruses.

    PubMed

    Juarez, Diana; Long, Kanya C; Aguilar, Patricia; Kochel, Tadeusz J; Halsey, Eric S

    2013-01-01

    Viruses from the Alphavirus genus are responsible for numerous arboviral diseases impacting human health throughout the world. Confirmation of acute alphavirus infection is based on viral isolation, identification of viral RNA, or a fourfold or greater increase in antibody titers between acute and convalescent samples. In convalescence, the specificity of antibodies to an alphavirus may be confirmed by plaque reduction neutralization test. To identify the best method for alphavirus and neutralizing antibody recognition, the standard solid method using a cell monolayer overlay with 0.4% agarose and the semisolid method using a cell suspension overlay with 0.6% carboxymethyl cellulose (CMC) overlay were evaluated. Mayaro virus, Una virus, Venezuelan equine encephalitis virus (VEEV), and Western equine encephalitis virus (WEEV) were selected to be tested by both methods. The results indicate that the solid method showed consistently greater sensitivity than the semisolid method. Also, a "semisolid-variant method" using a 0.6% CMC overlay on a cell monolayer was assayed for virus titration. This method provided the same sensitivity as the solid method for VEEV and also had greater sensitivity for WEEV titration. Modifications in plaque assay conditions affect significantly results and therefore evaluation of the performance of each new assay is needed.

  15. Reporter Immobilization Assay (REIA) for Bioconjugating Reactions.

    PubMed

    Schatte, Martin; Bocola, Marco; Roth, Teresa; Martinez, Ronny; Kopetzki, Erhard; Schwaneberg, Ulrich; Bönitz-Dulat, Mara

    2016-06-15

    Enzymes able to ligate biomolecules are emerging tools to generate site-specific bioconjugates. In this study we present a detection and screening method for bioconjugating enzymes which overcomes limitations of analytical methods such as HPLC or MS. These techniques are experimentally demanding and often limited in sensitivity and throughput compared to enzymatic assays. The principle of this Reporter Immobilization Assay (REIA) is the ligation of a reporter enzyme to a peptide carrying an affinity handle, which can be utilized for its isolation. The REIA system exhibits a high sensitivity with a linear range down to 1 μg/mL (55 nM), a variation coefficient of 6.5%, and can be performed cost-efficiently in 96-well microtiter plate format. The application of this assay allowed the characterization of a thiol transpeptidase sortase from S. aureus which is an important drug target and a biotechnological tool for ligation and modification of proteins. Thereby, yet-undetectable promiscuous activity of sortase could be detected, e.g., the acceptance of alanine as nucleophile. In addition, we were able to provide evidence that the REIA is suitable for high throughput screening of enzyme libraries using crude cellular extract with a throughput of 600 samples per hour. PMID:27182715

  16. Hyperpolarized NMR Probes for Biological Assays

    PubMed Central

    Meier, Sebastian; Jensen, Pernille R.; Karlsson, Magnus; Lerche, Mathilde H.

    2014-01-01

    During the last decade, the development of nuclear spin polarization enhanced (hyperpolarized) molecular probes has opened up new opportunities for studying the inner workings of living cells in real time. The hyperpolarized probes are produced ex situ, introduced into biological systems and detected with high sensitivity and contrast against background signals using high resolution NMR spectroscopy. A variety of natural, derivatized and designed hyperpolarized probes has emerged for diverse biological studies including assays of intracellular reaction progression, pathway kinetics, probe uptake and export, pH, redox state, reactive oxygen species, ion concentrations, drug efficacy or oncogenic signaling. These probes are readily used directly under natural conditions in biofluids and are often directly developed and optimized for cellular assays, thus leaving little doubt about their specificity and utility under biologically relevant conditions. Hyperpolarized molecular probes for biological NMR spectroscopy enable the unbiased detection of complex processes by virtue of the high spectral resolution, structural specificity and quantifiability of NMR signals. Here, we provide a survey of strategies used for the selection, design and use of hyperpolarized NMR probes in biological assays, and describe current limitations and developments. PMID:24441771

  17. Gamma neutron assay method and apparatus

    DOEpatents

    Cole, J.D.; Aryaeinejad, R.; Greenwood, R.C.

    1995-01-03

    The gamma neutron assay technique is an alternative method to standard safeguards techniques for the identification and assaying of special nuclear materials in a field or laboratory environment, as a tool for dismantlement and destruction of nuclear weapons, and to determine the isotopic ratios for a blend-down program on uranium. It is capable of determining the isotopic ratios of fissionable material from the spontaneous or induced fission of a sample to within approximately 0.5%. This is based upon the prompt coincidence relationships that occur in the fission process and the proton conservation and quasi-conservation of nuclear mass (A) that exists between the two fission fragments. The system is used in both passive (without an external neutron source) and active (with an external neutron source) mode. The apparatus consists of an array of neutron and gamma-ray detectors electronically connected to determine coincident events. The method can also be used to assay radioactive waste which contains fissile material, even in the presence of a high background radiation field. 7 figures.

  18. Transformation Assay for Identification of Psychrotrophic Achromobacters

    PubMed Central

    Juni, Elliot; Heym, Gloria A.

    1980-01-01

    The finding that many psychrotrophic, gram-negative, nonmotile, oxidase-positive coccobacilli (achromobacters) are competent for genetic transformation made possible the development of a transformation assay that permits recognition of genetically related strains. It has been demonstrated that 109 independently isolated achromobacters are genetically related since deoxyribonucleic acid samples from all of these organisms were able to transform a single competent auxotrophic strain to prototrophy. Genetically interacting bacteria included strains that lacked one or more of the characteristics typical for most achromobacters. An oxidase-negative mutant of one of these strains reacted positively in the transformation assay, unlike other oxidase-negative bacteria. Achromobacters were derived from fish, poultry, irradiated foods, seawater, and other sources. One strain previously classified as Micrococcus cryophilus has been shown to be related to the achromobacters. Two achromobacters had an optimum growth temperature of 35°C and behaved as typical mesophiles. The moraxellae and Acinetobacter were shown to be unrelated to the achromobacters by using the transformation assay. The ready demonstration of genetic relatedness provides a new basis for taxonomic grouping of the psychrotrophic achromobacters. Images PMID:16345673

  19. Hyperpolarized NMR probes for biological assays.

    PubMed

    Meier, Sebastian; Jensen, Pernille R; Karlsson, Magnus; Lerche, Mathilde H

    2014-01-16

    During the last decade, the development of nuclear spin polarization enhanced (hyperpolarized) molecular probes has opened up new opportunities for studying the inner workings of living cells in real time. The hyperpolarized probes are produced ex situ, introduced into biological systems and detected with high sensitivity and contrast against background signals using high resolution NMR spectroscopy. A variety of natural, derivatized and designed hyperpolarized probes has emerged for diverse biological studies including assays of intracellular reaction progression, pathway kinetics, probe uptake and export, pH, redox state, reactive oxygen species, ion concentrations, drug efficacy or oncogenic signaling. These probes are readily used directly under natural conditions in biofluids and are often directly developed and optimized for cellular assays, thus leaving little doubt about their specificity and utility under biologically relevant conditions. Hyperpolarized molecular probes for biological NMR spectroscopy enable the unbiased detection of complex processes by virtue of the high spectral resolution, structural specificity and quantifiability of NMR signals. Here, we provide a survey of strategies used for the selection, design and use of hyperpolarized NMR probes in biological assays, and describe current limitations and developments.

  20. The comet assay in human biomonitoring.

    PubMed

    Anderson, Diana; Dhawan, Alok; Laubenthal, Julian

    2013-01-01

    Human biomonitoring studies aim to identify potential exposures to environmental, occupational, or lifestyle toxicants in human populations and are commonly used by public health decision makers to predict disease risk. The Comet assay measures changes in genomic stability and is one of the most reliable biomarkers to indicate early biological effects, and therefore accepted by various governmental regulatory agencies. The appeal of the Comet assay lies in its relative simplicity, rapidity, sensitivity, and economic efficiency. Furthermore, the assay is known for its broad versatility, as it can be applied to virtually any human cell and easily adapted in order to detect particular biomarkers of interest, such as DNA repair capacity or single- and double-strand breaks. In a standard experiment, isolated single cells are first embedded in agarose, and then lysed in high-salt solutions in order to remove all cellular contents except the DNA attached to a nuclear scaffold. Subsequent electrophoresis results in accumulation of undamaged DNA sequences at the proximity of the nuclear scaffold, while damaged sequences migrate towards the anode. When visualized with fluorochromes, these migrated DNA fragments resemble a comet tail and can be quantified for their intensity and shape according to internationally drafted guidelines.

  1. Gamma neutron assay method and apparatus

    DOEpatents

    Cole, Jerald D.; Aryaeinejad, Rahmat; Greenwood, Reginald C.

    1995-01-01

    The gamma neutron assay technique is an alternative method to standard safeguards techniques for the identification and assaying of special nuclear materials in a field or laboratory environment, as a tool for dismantlement and destruction of nuclear weapons, and to determine the isotopic ratios for a blend-down program on uranium. It is capable of determining the isotopic ratios of fissionable material from the spontaneous or induced fission of a sample to within approximately 0.5%. This is based upon the prompt coincidence relationships that occur in the fission process and the proton conservation and quasi-conservation of nuclear mass (A) that exists between the two fission fragments. The system is used in both passive (without an external neutron source and active (with an external neutron source) mode. The apparatus consists of an array of neutron and gamma-ray detectors electronically connected to determine coincident events. The method can also be used to assay radioactive waste which contains fissile material, even in the presence of a high background radiation field.

  2. Quantitative comparisons of in vitro assays for estrogenic activities.

    PubMed Central

    Fang, H; Tong, W; Perkins, R; Soto, A M; Prechtl, N V; Sheehan, D M

    2000-01-01

    Substances that may act as estrogens show a broad chemical structural diversity. To thoroughly address the question of possible adverse estrogenic effects, reliable methods are needed to detect and identify the chemicals of these diverse structural classes. We compared three assays--in vitro estrogen receptor competitive binding assays (ER binding assays), yeast-based reporter gene assays (yeast assays), and the MCF-7 cell proliferation assay (E-SCREEN assay)--to determine their quantitative agreement in identifying structurally diverse estrogens. We examined assay performance for relative sensitivity, detection of active/inactive chemicals, and estrogen/antiestrogen activities. In this examination, we combined individual data sets in a specific, quantitative data mining exercise. Data sets for at least 29 chemicals from five laboratories were analyzed pair-wise by X-Y plots. The ER binding assay was a good predictor for the other two assay results when the antiestrogens were excluded (r(2) is 0.78 for the yeast assays and 0.85 for the E-SCREEN assays). Additionally, the examination strongly suggests that biologic information that is not apparent from any of the individual assays can be discovered by quantitative pair-wise comparisons among assays. Antiestrogens are identified as outliers in the ER binding/yeast assay, while complete antagonists are identified in the ER binding and E-SCREEN assays. Furthermore, the presence of outliers may be explained by different mechanisms that induce an endocrine response, different impurities in different batches of chemicals, different species sensitivity, or limitations of the assay techniques. Although these assays involve different levels of biologic complexity, the major conclusion is that they generally provided consistent information in quantitatively determining estrogenic activity for the five data sets examined. The results should provide guidance for expanded data mining examinations and the selection of appropriate

  3. Integrated air stream micromixer for performing bioanalytical assays on a plastic chip.

    PubMed

    Geissler, Matthias; Li, Kebin; Zhang, Xuefeng; Clime, Liviu; Robideau, Gregg P; Bilodeau, Guillaume J; Veres, Teodor

    2014-10-01

    This paper describes the design, functioning and use of an integrated mixer that relies on air flux to agitate microliter entities of fluid in an embedded microfluidic cavity. The system was fabricated from multiple layers of a thermoplastic elastomer and features circuits for both liquid and air supply along with pneumatic valves for process control. Internally-dyed polymer particles have been used to visualize flow within the fluid phase during agitation. Numerical modelling of the micromixer revealed an overall efficacy of 10(-1) to 10(-2) for momentum transfer at the air-water interface. Simulation of air vortex dynamics showed dependency of the flow pattern on the velocity of the flux entering the cavity. Three bioanalytical assays have been performed as proof-of-concept demonstrations. In a first assay, cells of Listeria monocytogenes were combined with magnetic nanoparticles (NPs), resulting in high-density coverage of the bacteria's surface with NPs after 1 min of agitation. This finding is contrasted by a control experiment without agitation for which interaction between bacteria and NPs remains low. In a second one, capture and release of genomic DNA from fungi through adsorption onto magnetic beads was tested and shown to be improved by agitation compared to non-agitated controls. A third assay finally involved fluorescently-labelled target oligonucleotide strands and polystyrene particles modified with DNA capture probes to perform detection of nucleic acids on beads. Excellent selectivity was obtained in a competitive hybridization process using a multiplexed micromixer chip design. PMID:25091476

  4. Magnetization dynamics using ultrashort magnetic field pulses

    NASA Astrophysics Data System (ADS)

    Tudosa, Ioan

    Very short and well shaped magnetic field pulses can be generated using ultra-relativistic electron bunches at Stanford Linear Accelerator. These fields of several Tesla with duration of several picoseconds are used to study the response of magnetic materials to a very short excitation. Precession of a magnetic moment by 90 degrees in a field of 1 Tesla takes about 10 picoseconds, so we explore the range of fast switching of the magnetization by precession. Our experiments are in a region of magnetic excitation that is not yet accessible by other methods. The current table top experiments can generate fields longer than 100 ps and with strength of 0.1 Tesla only. Two types of magnetic were used, magnetic recording media and model magnetic thin films. Information about the magnetization dynamics is extracted from the magnetic patterns generated by the magnetic field. The shape and size of these patterns are influenced by the dissipation of angular momentum involved in the switching process. The high-density recording media, both in-plane and perpendicular type, shows a pattern which indicates a high spin momentum dissipation. The perpendicular magnetic recording media was exposed to multiple magnetic field pulses. We observed an extended transition region between switched and non-switched areas indicating a stochastic switching behavior that cannot be explained by thermal fluctuations. The model films consist of very thin crystalline Fe films on GaAs. Even with these model films we see an enhanced dissipation compared to ferromagnetic resonance studies. The magnetic patterns show that damping increases with time and it is not a constant as usually assumed in the equation describing the magnetization dynamics. The simulation using the theory of spin-wave scattering explains only half of the observed damping. An important feature of this theory is that the spin dissipation is time dependent and depends on the large angle between the magnetization and the magnetic

  5. Nanoparticle-Based Histidine-Rich Protein-2 Assay for the Detection of the Malaria Parasite Plasmodium falciparum.

    PubMed

    Castro-Sesquen, Yagahira E; Kim, Chloe; Gilman, Robert H; Sullivan, David J; Searson, Peter C

    2016-08-01

    A nanoparticle-based assay for detection and quantification of Plasmodium falciparum histidine-rich protein 2 (HRP2) in urine and serum is reported. The assay uses magnetic beads conjugated with anti-HRP2 antibody for protein capture and concentration, and antibody-conjugated quantum dots for optical detection. Western blot analysis demonstrated that magnetic beads allow the concentration of HRP2 protein in urine by 20-fold. The concentration effect was achieved because large volume of urine can be incubated with beads, and magnetic separation can be easily performed in minutes to isolate beads containing HRP2 protein. Magnetic beads and quantum dots conjugated to anti-HRP2 antibodies allows the detection of low concentrations of HRP2 protein (0.5 ng/mL), and quantification in the range of 33-2,000 ng/mL corresponding to the range associated with non-severe to severe malaria. This assay can be easily adapted to a noninvasive point-of-care test for classification of severe malaria.

  6. Magnetism of Carbonados

    NASA Technical Reports Server (NTRS)

    Kletetschka, G.; Taylor, P. T.; Wasilewski, P. J.

    2000-01-01

    Origin of Carbonado is not clear. Magnetism of Carbonado comes from the surface, indicating contemporary formation of both the surface and magnetic carriers. The interior of carbonado is relatively free of magnetic phases.

  7. Samarium/Cobalt Magnets

    NASA Technical Reports Server (NTRS)

    Das, D.; Kumar, K.; Frost, R.; Chang, C.

    1985-01-01

    Intrinsic magnetic coercivities of samarium cobalt magnets made to approach theoretical limit of 350 kA/m by carefully eliminating oxygen from finished magnet by hot isostatic pressing (HIP). HIP process viable alternative to currently used sintering process.

  8. Magnetic Nanoparticle Quantitation with Low Frequency Magnetic Fields: Compensating for Relaxation Effects

    PubMed Central

    Weaver, John B.; Zhang, Xiaojuan; Kuehlert, Esra; Toraya-Brown, Seiko; Reeves, Daniel B.; Perreard, Irina M.; Fiering, Steven N.

    2013-01-01

    Quantifying the number of nanoparticles present in tissue is central to many in vivo and in vitro applications. Magnetic nanoparticles can be detected with high sensitivity both in vivo and in vitro using the harmonics of their magnetization produced in a sinusoidal magnetic field. However, relaxation effects damp the magnetic harmonics rendering them of limited use in quantitation. We show that an accurate measure of the number of nanoparticles can be made by correcting for relaxation effects. Correction for relaxation reduced errors of 50% for larger nanoparticles in high relaxation environments to 2%. The result is a method of nanoparticle quantitation capable of in vivo and in vitro applications including histopathology assays, quantitative imaging, drug delivery and thermal therapy preparation. PMID:23867287

  9. Magnetic particles

    NASA Technical Reports Server (NTRS)

    Chang, Manchium (Inventor); Colvin, Michael S. (Inventor); Rembaum, Alan (Inventor); Richards, Gil F. (Inventor)

    1987-01-01

    Metal oxide containing polymers and particularly styrene, acrylic or protein polymers containing fine, magnetic iron oxide particles are formed by combining a NO.sub.2 -substituted polymer with an acid such as hydrochloric acid in the presence of metal, particularly iron particles. The iron is oxidized to fine, black Fe.sub.3 O.sub.4 particles which deposit selectively on the polymer particles. Nitrated polymers are formed by reacting functionally substituted, nitrated organic compounds such as trinitrobenzene sulfonate or dinitrofluoro benzene with a functionally coreactive polymer such as an amine modified acrylic polymer or a protein. Other transition metals such as cobalt can also be incorporated into polymers using this method.

  10. Prospects for cellular mutational assays in human populations

    SciTech Connect

    Mendelsohn, M.L.

    1984-06-29

    Practical, sensitive, and effective human cellular assays for detecting somatic and germinal mutations would have great value in environmental mutagenesis and carcinogenesis studies. Such assays would fill the void between human mutagenicity and the data that exist from short-term tests and from mutagenicity in other species. This paper discusses the following possible human cellular assays: (1) HPRT (hypoxanthine phosphoribosyltransferase) somatic cell mutation based on 6-thioguanine resistance; (2) hemoglobin somatic cell mutation assay; (3) glycophorin somatic cell mutation assay; and (4) LDH-X sperm cell mutation assay. 18 references.

  11. Nanosecond magnetization reversal in nanocrystalline magnetic films

    NASA Astrophysics Data System (ADS)

    Rahman, I. Z.; Gandhi, A. A.; Khaddem-Mousavi, M. V.; Lynch, T. F.; Rahman, M. A.

    2007-03-01

    This paper reports on the investigation of dynamic magnetization reversal process in electrodeposited nanocrystalline Ni and Ni80Fe20 films by employing nanosecond magnetic pulse technique. The surface morphology has been investigated using SEM, EDAX, XRD and AFM analyses and static magnetic properties of the films are characterized by vibrating sample magnetometer (VSM). Two different techniques are designed and employed to study the nanosecond magnetization reversal process in nanocrystalline thin films: Magneto-Optical Kerr Effect (MOKE) and nanosecond pulsed field magnetometer. Results of dynamical behavior as a function of several variables such as magnitude of applied bias magnetic field, amplitude and width of the pulsed magnetic field are analyzed in detail using both techniques. A computer simulation package called Object Oriented Micro-Magnetic Framework (OOMMF) has been used to simulate the magnetic domain patterns of the samples.

  12. The Third Flight Magnet

    NASA Technical Reports Server (NTRS)

    McGhee, R. Wayne

    1998-01-01

    A self-shielded superconducting magnet was designed for the NASA Goddard Space Flight Center Adiabatic Demagnetization Refrigerator Program. This is the third magnet built from this design. The magnets utilize Cryomagnetics' patented ultra-low current technology. The magnetic system is capable of reaching a central field of two tesla at slightly under two amperes and has a total inductance of 1068 henries. This final report details the requirements of the magnet, the specifications of the resulting magnet, the test procedures and test result data for the third magnet (Serial # C-654-M), and recommended precautions for use of the magnet.

  13. Comparison of the luminescent ADP-Glo assay to a standard radiometric assay for measurement of protein kinase activity.

    PubMed

    Sanghera, Jasbinder; Li, Rick; Yan, Jun

    2009-12-01

    Many assay technologies have been developed and utilized to efficiently assay and screen against protein kinase targets. The radiometric assay format for assaying the protein kinase targets has been considered the "Gold Standard" format since it allows the direct readout of kinase functional activity and is a universal assay that is highly sensitive. However, the hazardous nature of the radiometric assay together with the regulatory hurdles has led to the development of alternative assay formats for assessing protein kinase activity measurements. The luminescent ADP-Glo assay has been developed as an alternative to radiometric format for assaying protein kinase targets. This assay allows the measurement of the ADP product formed during the kinase reaction. Therefore, the luminescent ADP-Glo assay is similar to the radiometric format in that it measures the direct product of the protein kinase reaction. Furthermore, since the ADP product is generated by all protein kinase reactions, this is a universal format that can be used for assaying any given protein kinase target. Analysis of data generated with multiple protein kinase targets and the luminescent ADP-Glo technology shows comparable results to the radiometric assay format. Therefore, the luminescent ADP-Glo assay is a robust new technology for evaluating catalytic function of protein kinases as well as other ATPases.

  14. A two-magnet strategy for improved mixing and capture from biofluids.

    PubMed

    Scherr, Thomas F; Ryskoski, Hayley B; Doyle, Andrew B; Haselton, Frederick R

    2016-03-01

    Magnetic beads are a popular method for concentrating biomolecules from solution and have been more recently used in multistep pre-arrayed microfluidic cartridges. Typical processing strategies rely on a single magnet, resulting in a tight cluster of beads and requiring long incubation times to achieve high capture efficiencies, especially in highly viscous patient samples. This report describes a two-magnet strategy to improve the interaction of the bead surface with the surrounding fluid inside of a pre-arrayed, self-contained assay-in-a-tube. In the two-magnet system, target biomarker capture occurs at a rate three times faster than the single-magnet system. In clinically relevant biomatrices, we find a 2.5-fold improvement in biomarker capture at lower sample viscosities with the two-magnet system. In addition, we observe a 20% increase in the amount of protein captured at high viscosity for the two-magnet configuration relative to the single magnet approach. The two-magnet approach offers a means to achieve higher biomolecule extraction yields and shorter assay times in magnetic capture assays and in self-contained processor designs. PMID:27158286

  15. Electrochemical chip-based genomagnetic assay for detection of high-risk human papillomavirus DNA.

    PubMed

    Bartosik, Martin; Durikova, Helena; Vojtesek, Borivoj; Anton, Milan; Jandakova, Eva; Hrstka, Roman

    2016-09-15

    Cervical cancer, being the fourth leading cause of cancer death in women worldwide, predominantly originates from a persistent infection with a high-risk human papillomavirus (HPV). Detection of DNA sequences from these high-risk strains, mostly HPV-16 and HPV-18, represents promising strategy for early screening, which would help to identify women with higher risk of cervical cancer. In developing countries, inadequate screening options lead to disproportionately high mortality rates, making a fast and inexpensive detection schemes highly important. Electrochemical sensors and assays offer an alternative to current methods of detection. We developed an electrochemical-chip based assay, in which target HPV DNA is captured via magnetic bead-modified DNA probes, followed by an antidigoxigenin-peroxidase detection system at screen-printed carbon electrode chips, enabling parallel measurements of eight samples simultaneously. We show sensitive detection in attomoles of HPV DNA, selective discrimination between HPV-16 and HPV-18 and good reproducibility. Most importantly, we show application of the assay into both cancer cell lines and cervical smears from patients. The electrochemical results correlated well with standard methods, making this assay potentially applicable in clinical practice.

  16. Nondestructive boxed transuranic (TRU) waste assay systems

    NASA Astrophysics Data System (ADS)

    Caldwell, John T.; Jones, Stephanie A.; Lucero, Randy F.

    1999-01-01

    A brief history of boxed waste assay systems (primarily those developed at Los Alamos National Laboratory) is presented. The characteristics and design process involved with current generation systems--as practiced by BII--are also discussed in some detail. Finally, a specific boxed waste assay system and acceptance test results are presented. This system was developed by BII and installed at the Waste Receiving and Packaging (WRAP) facility in Hanford, Washington in early 1997. The WRAP system combines imaging passive/active neutron (IPAN) techniques with gamma- ray energy analysis (GEA) to assay crates up to 2.5 m X 2.5 m X 6.5 m in size. (Systems that incorporate both these methodologies are usually denoted IPAN/GEA types.) Two separate gamma-ray measurements are accomplished utilizing 16 arrayed NaI detectors and a moveable HPGe detector, while 3He detectors acquire both active and passive neutron data. These neutron measurements use BII's proprietary imaging methodology. Acceptance testing of the system was conducted at Hanford in January 1998. The system's operating performance was evaluated based on accuracy and sensitivity requirements for three different matrix types. Test results indicate an average 13% active mode accuracy for 10 nCi/g loadings of Pu waste and 5% passive mode accuracy for 10 g loadings of Pu waste. Sensitivity testing demonstrated an active mode lower limit of detection of less than 5 nCi/g of 239Pu for the medium matrix and less than 20 pCi/g of fission and activation products at 3(sigma) above background.

  17. Nondestructive assay measurements of GNEP related materials

    SciTech Connect

    Santi, Peter A; Crooks, William J.; Geist, William H.; Gonzales, Robert; Helland, Carolyn A.; Jackson, Jay M.; Frame, Katherine C.; Martinez, Michael M.; Scherer, Caroylnn P.; Vo, Duc T.

    2008-06-12

    Because the reprocessing technologies that are currently being considered for the Global Nuclear Energy Partnership (GNEP) will keep various actinides commingled with plutonium at all times throughout the process, the resulting nuclear fuel that is intended for the Advanced Burner Reactor will present unique measurement challenges for the various Nondestructive Assay (NDA) techniques. In order to begin clarifying which types of materials and measurement scenarios that may exist within GNEP require the development of new measurement technologies, an initial series of measurements have been performed on materials with radiation properties that are similar to those being considered within GNEP.

  18. A universal nanoparticle cell secretion capture assay.

    PubMed

    Fitzgerald, Wendy; Grivel, Jean-Charles

    2013-02-01

    Secreted proteins play an important role in intercellular interactions, especially between cells of the immune system. Currently, there is no universal assay that allows a simple noninvasive identification and isolation of cells based on their secretion of various products. We have developed such a method. Our method is based on the targeting, to the cell surface, of heterofunctional nanoparticles coupled to a cell surface-specific antibody and to a secreted protein-specific antibody, which captures the secreted protein on the surface of the producing cell. Importantly, this method does not compromise cellviability and is compatible with further culture and expansion of the secreting cells.

  19. Assay of the Martian Regolith with Neutrons

    NASA Technical Reports Server (NTRS)

    Drake, Darrell M.; Reedy, R.; Jakowsky, B.; Clark, B.; Squyres, S.

    1998-01-01

    Different aspects of assaying Martian regolith using neutrons have been investigated. The epithermal portion of moderated neutrons spectra is dramatically effected by the presence of hydrogen (usually in the form of water). A simple analytic formula has been derived to describe the amplitude of this portion of the neutron spectrum as a function of water concentration. Several demonstration experiments have been performed and modeled with a Monte Carlo code. Results of these experiments generally agreed with the calculations to within 20%. In addition to He-3 detectors, lithium-glass scintillators and U-238 fission ion chambers were investigated to determine their applicability to space experiments.

  20. Test procedure for boxed waste assay system

    SciTech Connect

    Wachter, J.

    1994-12-07

    This document, prepared by Los Alamos National Laboratory`s NMT-4 group, details the test methodology and requirements for Acceptance/Qualification testing of a Boxed Waste Assay System (BWAS) designed and constructed by Pajarito Scientific Corporation. Testing of the BWAS at the Plutonium Facility (TA55) at Los Alamos National Laboratory will be performed to ascertain system adherence to procurement specification requirements. The test program shall include demonstration of conveyor handling capabilities, gamma ray energy analysis, and imaging passive/active neutron accuracy and sensitivity. Integral to these functions is the system`s embedded operating and data reduction software.

  1. Protein kinase profiling assays: a technology review.

    PubMed

    Wang, Yuren; Ma, Haiching

    2015-11-01

    Protein kinases have become one of the most intensively pursued classes of drug targets for many diseases such as cancers and inflammatory diseases. Kinase profiling work seeks to understand general selectivity trends of lead compounds across the kinome, which help with target selection, compound prioritization, and potential implications in toxicity. Under the current drug discovery process, screening of compounds against comprehensive panels of kinases and their mutants has become the standard approach. Many screening assays and technologies which are compatible for high-throughput screening (HTS) against kinases have been extensively pursued and developed.

  2. Lipase turbidimetric assay and acute pancreatitis.

    PubMed

    Orda, R; Orda, S; Baron, J; Wiznitzer, T

    1984-04-01

    The simplified turbidimetric assay for lipase activity was used for the differential diagnosis of acute pancreatitis. Serum lipase levels were found to be increased in a group of 17 patients in whom acute pancreatitis was clinically suspected and confirmed by a high ACCR and decreased uptake of the radionuclide in the pancreas scan. The lipase levels were within normal limits in a control group of 14 patients suffering from diseases other than acute pancreatitis. The turbidimetric test was helpful for rapid quantitative determination of serum lipase and thus for the early and accurate diagnosis of acute pancreatitis. PMID:6200277

  3. Assaying the Potency of Influenza Vaccines

    PubMed Central

    Minor, Philip D.

    2015-01-01

    The potency of vaccines must be determined to ensure that the appropriate dose is given. The manufacture and assessment of influenza vaccines are complicated by the continuously changing nature of the pathogen, which makes efficacy estimates difficult but also confounds attempts to produce a well-validated, consistent potency assay. Single radial diffusion has been used for decades and provides a relatively simple way to measure the amount of biologically active materials present in the vaccine. It requires reagents, which are updated on a regular, frequently yearly, basis and alternative methods continue to be sought. PMID:26344948

  4. Electrochemical and Magnetic Technologies for Bio Applications

    NASA Astrophysics Data System (ADS)

    Matsunaga, Tadashi; Tanaka, Tsuyoshi

    The electrochemical and magnetic biosensors have an advantage because of the easy miniaturization of electric device components as compared with photometric instruments. These technologies have been applied to develop portable, compact and inexpensive biochip devices. A commercially successful example is the glucose sensor using enzyme transducers, which was originally reported by Clark and Lyons [1] to measure glucose by detecting the decrease in oxygen by pO2 electrode when glucose is converted to gluconic acid and hydrogen peroxide. Electrochemical biosensors can be separated into three typical assay systems using amperometric, potentiometric or conductometric transducers. Furthermore, various magnetosensors using magnetic particles have been developed over a decade in place of photometric biosensors. In this chapter, recent advances in electrochemical and magnetic biosensors toward development of portable, compact and inexpensive biochip devices have been focused.

  5. Magnetic liposomes for colorectal cancer cells therapy by high-frequency magnetic field treatment

    NASA Astrophysics Data System (ADS)

    Hardiansyah, Andri; Huang, Li-Ying; Yang, Ming-Chien; Liu, Ting-Yu; Tsai, Sung-Chen; Yang, Chih-Yung; Kuo, Chih-Yu; Chan, Tzu-Yi; Zou, Hui-Ming; Lian, Wei-Nan; Lin, Chi-Hung

    2014-09-01

    In this study, we developed the cancer treatment through the combination of chemotherapy and thermotherapy using doxorubicin-loaded magnetic liposomes. The citric acid-coated magnetic nanoparticles (CAMNP, ca. 10 nm) and doxorubicin were encapsulated into the liposome (HSPC/DSPE/cholesterol = 12.5:1:8.25) by rotary evaporation and ultrasonication process. The resultant magnetic liposomes ( ca. 90 to 130 nm) were subject to characterization including transmission electron microscopy (TEM), dynamic light scattering (DLS), X-ray diffraction (XRD), zeta potential, Fourier transform infrared (FTIR) spectrophotometer, and fluorescence microscope. In vitro cytotoxicity of the drug carrier platform was investigated through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay using L-929 cells, as the mammalian cell model. In vitro cytotoxicity and hyperthermia (inductive heating) studies were evaluated against colorectal cancer (CT-26 cells) with high-frequency magnetic field (HFMF) exposure. MTT assay revealed that these drug carriers exhibited no cytotoxicity against L-929 cells, suggesting excellent biocompatibility. When the magnetic liposomes with 1 μM doxorubicin was used to treat CT-26 cells in combination with HFMF exposure, approximately 56% cells were killed and found to be more effective than either hyperthermia or chemotherapy treatment individually. Therefore, these results show that the synergistic effects between chemotherapy (drug-controlled release) and hyperthermia increase the capability to kill cancer cells.

  6. Magnetic liposomes for colorectal cancer cells therapy by high-frequency magnetic field treatment

    PubMed Central

    2014-01-01

    In this study, we developed the cancer treatment through the combination of chemotherapy and thermotherapy using doxorubicin-loaded magnetic liposomes. The citric acid-coated magnetic nanoparticles (CAMNP, ca. 10 nm) and doxorubicin were encapsulated into the liposome (HSPC/DSPE/cholesterol = 12.5:1:8.25) by rotary evaporation and ultrasonication process. The resultant magnetic liposomes (ca. 90 to 130 nm) were subject to characterization including transmission electron microscopy (TEM), dynamic light scattering (DLS), X-ray diffraction (XRD), zeta potential, Fourier transform infrared (FTIR) spectrophotometer, and fluorescence microscope. In vitro cytotoxicity of the drug carrier platform was investigated through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay using L-929 cells, as the mammalian cell model. In vitro cytotoxicity and hyperthermia (inductive heating) studies were evaluated against colorectal cancer (CT-26 cells) with high-frequency magnetic field (HFMF) exposure. MTT assay revealed that these drug carriers exhibited no cytotoxicity against L-929 cells, suggesting excellent biocompatibility. When the magnetic liposomes with 1 μM doxorubicin was used to treat CT-26 cells in combination with HFMF exposure, approximately 56% cells were killed and found to be more effective than either hyperthermia or chemotherapy treatment individually. Therefore, these results show that the synergistic effects between chemotherapy (drug-controlled release) and hyperthermia increase the capability to kill cancer cells. PMID:25246875

  7. HPLC-MTT assay: anticancer activity of aqueous garlic extract is from allicin.

    PubMed

    Lee, Jenny; Gupta, Shalini; Huang, Jin-Sheng; Jayathilaka, Lasanthi P; Lee, Bao-Shiang

    2013-05-15

    A strategy using reversed-phase high-performance liquid chromatography (HPLC), thin layer chromatography (TLC), mass spectrometry (MS), nuclear magnetic resonance (NMR), chemical synthesis, and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) cell viability assay to identify allicin as the active anticancer compound in aqueous garlic extract (AGE) is described. Changing the pH of AGE from 7.0 to 5.0 eliminated interfering molecules and enabled a clean HPLC separation of the constituents in AGE. MTT assay of the HPLC fractions identified an active fraction. Further analysis by TLC, MS, and NMR verified the active HPLC fraction as allicin. Chemically synthesized allicin was used to provide further confirmation. The results clearly identify the active compound in AGE as allicin.

  8. Enzyme-linked fluorescence assay: Ultrasensitive solid-phase assay for detection of human rotavirus.

    PubMed Central

    Yolken, R H; Stopa, P J

    1979-01-01

    Enzyme-linked immunosorbent assay (ELISA) has proven to be a useful assay system for the direct detection of infectious agents. However, when the usual color-producing substrates are employed, relatively large amounts of substrate must be hydrolyzed by the bound enzyme before detection can be achieved. We attempted to improve the sensitivity of ELISA by utilizing a substrate that yields a fluorescent product on enzyme action. The enzyme-linked fluorescence assay (ELFA) based on this principle was approximately 100 times more sensitive than the corresponding ELISA or radioimmunoassay for the detection of human rotavirus in a standard stool suspension. In addition, the ELFA for human rotavirus was capable of detecting antigen in six specimens that were negative by ELISA. Five of these specimens were obtained late in the course of confirmed rotavirus infections. ELFA provides a simple, reliable, ultrasensitive method for the rapid detection of viral antigen. PMID:226564

  9. Clinical Assay Development Support - Office of Cancer Clinical Proteomics Research

    Cancer.gov

    The NCI’s Division of Cancer Treatment and Diagnosis and the Cancer Diagnosis Program announce a request for applications for the Clinical Assay Development Program (CADP) for investigators seeking clinical assay development and validation resources.

  10. A Spectrophotometric Assay Optimizing Conditions for Pepsin Activity.

    ERIC Educational Resources Information Center

    Harding, Ethelynda E.; Kimsey, R. Scott

    1998-01-01

    Describes a laboratory protocol optimizing the conditions for the assay of pepsin activity using the Coomasie Blue dye binding assay of protein concentration. The dye bonds through strong, noncovalent interactions to basic and aromatic amino acid residues. (DDR)

  11. Stress Response to High Magnetic Fields in Transgenic Arabidopsis thaliana Plants.

    NASA Astrophysics Data System (ADS)

    Morgan, A. N.; Watson, B. C.; Maloney, J. R.; Meisel, M. W.; Brooks, J. S.; Paul, A.-L.; Ferl, R. J.

    2000-03-01

    With increasingly greater strength magnetic fields becoming available in research and medicine, the response of living tissue exposed to high magnetic fields has come under investigation. In this experiment, genetically engineered arabidopsis plants were exposed to homogeneous magnetic fields of varying strengths using a superconducting NMR magnet (0 to 9 T) at UF and a resistive magnet (0 to 25 T) at the NHMFL. The engineered plants produce the enzyme β-glucaronidase (GUS) when under stressful environmental conditions. The level of GUS activity is determined through qualitative histochemical assays and quantitative fluorometric assays. The control group of plants experienced baseline levels of GUS activity, but some of the plants that were exposed to magnetic fields in excess of 9 T show increased stress response. Additional information is available at http://www.phys.ufl.edu/ ~meisel/maglev.htm.

  12. Magnetically Responsive Negative Acoustic Contrast Microparticles for Bioanalytical Applications.

    PubMed

    Ohiri, Korine A; Evans, Benjamin A; Shields, C Wyatt; Gutiérrez, Robert A; Carroll, Nick J; Yellen, Benjamin B; López, Gabriel P

    2016-09-28

    Smart colloidal particles are routinely used as carriers for biological molecules, fluorescent reporters, cells, and other analytes for the purposes of sample preparation and detection. However, such particles are typically engineered to respond to a single type of stimulus (e.g., commercial magnetic beads to magnetic fields). Here, we demonstrate a unique class of particles that display both positive magnetic contrast and negative acoustic contrast in water. This dual functionality allows for fine spatiotemporal control, enabling multiple separation modalities and increasing the utility of the particles in various chemical and biological assays.

  13. Magnetically Responsive Negative Acoustic Contrast Microparticles for Bioanalytical Applications.

    PubMed

    Ohiri, Korine A; Evans, Benjamin A; Shields, C Wyatt; Gutiérrez, Robert A; Carroll, Nick J; Yellen, Benjamin B; López, Gabriel P

    2016-09-28

    Smart colloidal particles are routinely used as carriers for biological molecules, fluorescent reporters, cells, and other analytes for the purposes of sample preparation and detection. However, such particles are typically engineered to respond to a single type of stimulus (e.g., commercial magnetic beads to magnetic fields). Here, we demonstrate a unique class of particles that display both positive magnetic contrast and negative acoustic contrast in water. This dual functionality allows for fine spatiotemporal control, enabling multiple separation modalities and increasing the utility of the particles in various chemical and biological assays. PMID:27622731

  14. Storage and Assay of Tritium in STAR

    SciTech Connect

    Glen R. Longhurst; Robert A. Anderl; Robert J. Pawelko

    2004-09-01

    The Safety and Tritium Applied Research (STAR) facility has recently been commissioned to investigate tritium-related safety questions for fusion and other technologies. The authorized inventory of tritium is 1.6 grams, the threshold quantity for nuclear facility classification. A key capability in successful operation of the STAR facility is the ability to receive, inventory, and dispense tritium to the various experiments underway there. The system central to that function is the Tritium Storage and Assay System (SAS). The SAS has four major functions: (1) receiving and holding tritium from shipping containers brought into the STAR facility, (2) assaying the amount of tritium in the SAS, (3) dispensing tritium to secondary beds or containers used for transferring it to the experimental systems in the STAR facility, and (4) purifying hydrogen isotopes from non-hydrogen species. To that may be added a fifth, optional function, isotopic separation of hydrogen isotopes using bed-to-bed transfer techniques. This paper documents the design and operation of the STAR SAS and the procedures used for tritium accountancy in the STAR facility.

  15. Rapid screening assay for calcium bioavailability studies

    SciTech Connect

    Luhrsen, K.R.; Hudepohl, G.R.; Smith, K.T.

    1986-03-01

    Calcium bioavailability has been studied by numerous techniques. The authors report here the use of the gamma emitting isotope of calcium (/sup 47/Ca) in a whole body retention assay system. In this system, calcium sources are administered by oral gavage and subsequent counts are determined and corrected for isotopic decay. Unlike iron and zinc retention curves, which exhibit a 2-3 day equilibration period, calcium reaches equilibration after 24 hours. Autoradiographic analysis of the femurs indicate that the newly absorbed calcium is rapidly distributed to the skeletal system. Moreover, the isotope is distributed along the entire bone. Comparisons of calcium bioavailability were made using intrinsic/extrinsic labeled milk from two species i.e. rat and goat as well as CaCO/sub 3/. In addition, extrinsic labeled cow milk was examined. In the rat, the extrinsic labeled calcium from milk was better absorbed than the intrinsic calcium. This was not the case in goat milk or the calcium carbonate which exhibited no significant differences. Chromatographic analysis of the labeled milk indicates a difference in distribution of the /sup 47/Ca. From these data, the authors recommend the use of this assay system in calcium bioavailability studies. The labeling studies and comparisons indicate caution should be used, however, in labeling techniques and species milk comparison.

  16. The clinical value of tissue factor assays.

    PubMed

    Francis, J L; Carvalho, M; Francis, D A

    1995-06-01

    Tissue factor (TF) is now considered to be the primary physiologic activator of the blood coagulation system. Coupled with recent advances in our understanding of the biochemistry of TF this has heightened interest in measuring aspects of TF activity in disease states. Expression of TF by blood monocytes in various diseases is an established trigger for intravascular coagulation and there is now a considerable body of experience with its measurement. This has considerable clinical potential although more widespread application awaits a consensus on the most appropriate methodologic approach to its measurement. TF can be detected in urine and may reflect the activation state of renal macrophages. Urinary TF is increased in cancer and could have diagnostic and prognostic value in a variety of malignant diseases. Finally, it is now possible to measure soluble TF in plasma. One such assay is commercially available and is technically simple to perform. The clinical value of such assays, however, must await better understanding of the source and function of soluble TF in plasma.

  17. Application of nondestructive assay techniques in Kazakstan

    SciTech Connect

    Sprinkle, J.K. Jr.; Butler, G.; Collins, M.

    1997-11-01

    As Kazakstan has transitioned from being part of the Soviet Union to a nonweapons state (Treaty of Nonproliferation of Nuclear Weapons [NPT] signatory) under International Atomic Energy Agency (IAEA) inspections, significant changes have been required. Some of these changes have occurred in nuclear material protection, control, and accounting at the four nuclear facility sites in the Republic of Kazakstan. Specifically, the Republic of Kazakstan has changed from relying primarily on a subset of physical protection methods to a graded safeguards approach using a balance of material control, material accounting, and physical protection. Once more intensive material control and accounting procedures and systems are in place, a necessary step is to supply the accounting systems with measured values of high quality. This need can be met with destructive and nondestructive methods. Material control systems can also use qualitative nondestructive assay information as input. This paper will discuss the nondestructive assay techniques and systems the US Department of Energy (DOE) is providing to Kazakstan under both DOE programs and the Cooperative Threat Reduction Act as part of the nuclear material control and accounting upgrades at four facilities in Kazakstan. 4 refs., 6 figs.

  18. The clinical value of tissue factor assays.

    PubMed

    Francis, J L; Carvalho, M; Francis, D A

    1995-06-01

    Tissue factor (TF) is now considered to be the primary physiologic activator of the blood coagulation system. Coupled with recent advances in our understanding of the biochemistry of TF this has heightened interest in measuring aspects of TF activity in disease states. Expression of TF by blood monocytes in various diseases is an established trigger for intravascular coagulation and there is now a considerable body of experience with its measurement. This has considerable clinical potential although more widespread application awaits a consensus on the most appropriate methodologic approach to its measurement. TF can be detected in urine and may reflect the activation state of renal macrophages. Urinary TF is increased in cancer and could have diagnostic and prognostic value in a variety of malignant diseases. Finally, it is now possible to measure soluble TF in plasma. One such assay is commercially available and is technically simple to perform. The clinical value of such assays, however, must await better understanding of the source and function of soluble TF in plasma. PMID:7647219

  19. Standardization of anti-DNA antibody assays.

    PubMed

    Pisetsky, David S

    2013-07-01

    Antibodies to DNA (anti-DNA) are the serological hallmark of systemic lupus erythematosus and represent important biomarkers for clinical and research purposes. These antibodies are part of a family of antibodies to nucleosomes and bind to conserved sites widely present on DNA. While the value of anti-DNA as a biomarker is well established, the assay for these antibodies has involved a variety of DNA sources and systems to detect DNA-anti-DNA interactions. The influence of these variations on antibody detection has complicated assay standardization. As an antigen, DNA has unique features since it is a highly charged polymer that has structural heterogeneity. This heterogeneity can affect antigenicity which can vary on the basis of DNA origin, size, conformation and mobility. In addition, as a polymer, DNA can promote patterns of antibody binding based on monogamous or bivalent interaction which require an extended polynucleotide structure. Understanding the nature of DNA as an antigen can facilitate interpretation of serological tests and underpin efforts at better standardization.

  20. Assaying Environmental Nickel Toxicity Using Model Nematodes

    PubMed Central

    Rudel, David; Douglas, Chandler D.; Huffnagle, Ian M.; Besser, John M.; Ingersoll, Christopher G.

    2013-01-01

    Although nickel exposure results in allergic reactions, respiratory conditions, and cancer in humans and rodents, the ramifications of excess nickel in the environment for animal and human health remain largely undescribed. Nickel and other cationic metals travel through waterways and bind to soils and sediments. To evaluate the potential toxic effects of nickel at environmental contaminant levels (8.9-7,600 µg Ni/g dry weight of sediment and 50-800 µg NiCl2/L of water), we conducted assays using two cosmopolitan nematodes, Caenorhabditis elegans and Pristionchus pacificus. We assayed the effects of both sediment-bound and aqueous nickel upon animal growth, developmental survival, lifespan, and fecundity. Uncontaminated sediments were collected from sites in the Midwestern United States and spiked with a range of nickel concentrations. We found that nickel-spiked sediment substantially impairs both survival from larval to adult stages and adult longevity in a concentration-dependent manner. Further, while aqueous nickel showed no adverse effects on either survivorship or longevity, we observed a significant decrease in fecundity, indicating that aqueous nickel could have a negative impact on nematode physiology. Intriguingly, C. elegans and P. pacificus exhibit similar, but not identical, responses to nickel exposure. Moreover, P. pacificus could be tested successfully in sediments inhospitable to C. elegans. Our results add to a growing body of literature documenting the impact of nickel on animal physiology, and suggest that environmental toxicological studies could gain an advantage by widening their repertoire of nematode species. PMID:24116204

  1. Rapid Multiple Immunoenzyme Assay of Mycotoxins

    PubMed Central

    Urusov, Alexandr E.; Zherdev, Anatoly V.; Petrakova, Alina V.; Sadykhov, Elchin G.; Koroleva, Olga V.; Dzantiev, Boris B.

    2015-01-01

    Mycotoxins are low molecular weight fungal metabolites that pose a threat as toxic contaminants of food products, thereby necessitating their effective monitoring and control. Microplate ELISA can be used for this purpose, but this method is characteristically time consuming, with a duration extending to several hours. This report proposes a variant of the ELISA method for the detection and quantification of three mycotoxins, ochratoxin A, aflatoxin B1 and zearalenone, in the kinetic regime. The main requirement for the proposed kinetic protocol was to provide a rapid method that combined sensitivity and accuracy. The use of biotin with an extended spacer together with a streptavidin–polyperoxidase conjugate provided high signal levels, despite these interactions occurring under non-equilibrium conditions. Duration of the individual mycotoxin assays was 20 min, whereas the analysis of all three mycotoxins in parallel reached a maximum duration of 25 min. Recovery of at least 95% mycotoxins in water-organic extracts was shown. The developed assays were successfully validated using poultry processing products and corn samples spiked with known quantities of mycotoxins. The detection limits for aflatoxin B1, ochratoxin A and zearalenone in these substances were 0.24, 1.2 and 3 ng/g, respectively. PMID:25633750

  2. Improved flow cytometer measurement of binding assays

    NASA Astrophysics Data System (ADS)

    Saunders, G. C.

    1984-05-01

    A method of measuring binding assays is carried out with different size particles wherein the binding assay sample is run through a flow cytometer without separating the sample from the marking agent. The amount of a binding reactant present in a sample is determined by providing particles with a coating of binder and also known quantity of smaller particles with a coating of binder reactant. The smaller particles also contain a fluorescent chemical. The particles are combined with the sample and the binding reaction is allowed to occur for a set length of time followed by combining the smaller particles with the mixture of the particles and the sample produced and allowing the binding reactions to proceed to equilibrium. The fluorescence and light scatter of the combined mixture is then measured as the combined mixture passes through a flow cytometer equipped with a laser to bring about fluorescence, and the number of fluorescent events are compared. A similar method is also provided for determining the amount of antigen present in the sample by providing spheres with an antibody coating and some smaller spheres with an antigen coating.

  3. Preliminary assay reveals quality of Mars blend

    SciTech Connect

    Rhodes, A.K.

    1996-06-10

    Shell Oil Co. has assayed a preproduction well sample of Mars blend, from the Louisiana Gulf of Mexico. The assay shows the crude to have an API gravity of 31{degree} and a sulfur content of 2.0 wt%. Distillation data for the crude are shown in this paper. When partners Shell (71.5%) and BP (28.5%) begin commercial production in July, several producing zones will be tapped through a number of wells. Shell says the final sales quality of Mars Blend may vary from the qualities of this representative sample. The crude will be produced from a tension-leg platform (TLP) at a water depth of 2,940 ft (OGJ, Apr. 8, p. 23). The TLP is anchored in Mississippi Canyon block 807, about 130 miles southwest of New Orleans (map). Mars field will be the principal source of the crude initially. Shell says production will escalate to about 100,000 b/d crude and 110 MMcfd natural gas in 1997. Ultimate recovery is estimated to be 700 million BOE.

  4. Standardisation of the factor H autoantibody assay.

    PubMed

    Watson, Rachael; Lindner, Susanne; Bordereau, Pauline; Hunze, Eva-Maria; Tak, Federico; Ngo, Stéphanie; Zipfel, Peter F; Skerka, Christine; Dragon-Durey, Marie-Agnes; Marchbank, Kevin J

    2014-01-01

    The screening of all atypical haemolytic uraemic syndrome (aHUS) patients for factor H autoantibodies is best practice. However, there is no consensus assay for the reporting of factor H autoantibody titres. In this study, three European complement laboratories with expertise in the field of autoantibody testing address this by systematically evaluating several ELISA methods used for the detection of factor H autoantibodies. All methods tested adequately detect high titre samples. However, this study recommends the Paris method for the detection and reporting of factor H autoantibodies to be used when setting up a factor H autoantibody screen. The importance of individual sample background subtraction in these ELISA tests was established. The use of a relative or arbitrary unit index with a common positive and negative serum allowed for consistent comparison of findings from different test centres. Therefore, it is recommended that a standard arbitrary unit scale based on a titration curve from a common positive anti-serum be adopted to allow future establishment of the relative importance of particular titres of factor H autoantibodies in aHUS. Systematic assay for the presence of factor H autoantibodies in patients using the Paris method will provide the longitudinal analysis needed to fully establish the importance of factor H autoantibodies in disease. This will feed into additional research to clarify whether additional factors have a bearing on the phenotype/outcome of autoimmune aHUS. PMID:23891327

  5. Improved flow cytometer measurement of binding assays

    DOEpatents

    Saunders, G.C.

    1984-05-30

    The invention relates to a method of measuring binding assays carried out with different size particles wherein the binding assay sample is run through a flow cytometer without separating the sample from the marking agent. The amount of a binding reactant present in a sample is determined by providing particles with a coating of binder and also a known quantity of smaller particles with a coating of binder reactant. The binding reactant is the same as the binding reactant present in the sample. The smaller particles also contain a fluorescent chemical. The particles are combined with the sample and the binding reaction is allowed to occur for a set length of time followed by combining the smaller particles with the mixture of the particles and the sample produced and allowing the binding reactions to proceed to equilibrium. The fluorescence and light scatter of the combined mixture is then measured as the combined mixture passes through a flow cytometer equipped with a laser to bring about fluorescence, and the number and strength of fluorescent events are compared. A similar method is also provided for determining the amount of antigen present in the sample by providing spheres with an antibody coating and some smaller spheres with an antigen coating. (LEW)

  6. Survey of assay methods of antivenins

    PubMed Central

    Grasset, E.

    1957-01-01

    In view of the multiplicity of methods used at present for the preparation and assay of antivenins and as a first step towards the international standardization of antivenins, it seemed advisable to make a comparative study of the methods used in the institutes specializing in the production of these sera. With this end in view, the author circulated to the serologists of institutes concerned a detailed questionnaire on the assay methods used for the determination of the neutralization potency of the various types of antivenins prepared under their direction. The information supplied by these institutes is reproduced, in condensed form, in this report and is analysed by the author. The author emphasizes that the great variety in the constitution of venoms necessitates: (1) the use of monovalent standard sera against homologous “test” venoms of high activity and stability; and (2) the establishment, on a regional basis, of standard antivenins corresponding to groups of snakes characterized by venoms of common or closely related antigenic constitution. PMID:13413648

  7. Synthesis and Assays of Inhibitors of Methyltransferases.

    PubMed

    Cai, X-C; Kapilashrami, K; Luo, M

    2016-01-01

    Epigenetic regulation requires site-specific modification of the genome and is involved in multiple physiological processes and disease etiology. Methyltransferases, which catalyze the transfer of a methyl group from S-adenosyl-l-methionine (SAM) to various substrates, are critical components of the epigenetic machinery. This group of enzymes can methylate diverse substrates including DNA, RNA, proteins, and small-molecule metabolites. Their dysregulation has also been implicated in multiple disease states such as cancer, neurological, and cardiovascular disorders. Developing potent and selective small-molecule inhibitors of methyltransferases is valuable not only for therapeutic intervention but also for investigating the roles of these enzymes in disease progression. In this chapter, we will discuss the strategies of designing and synthesizing methyltransferases inhibitors based on the SAM scaffold. Following the section of inhibitor design, we will briefly review representative assays that are available to evaluate the potency of these inhibitors along with a detailed description of the most commonly used radiometric assay. PMID:27423865

  8. Assaying environmental nickel toxicity using model nematodes

    USGS Publications Warehouse

    Rudel, David; Douglas, Chandler; Huffnagle, Ian; Besser, John M.; Ingersoll, Christopher G.

    2013-01-01

    Although nickel exposure results in allergic reactions, respiratory conditions, and cancer in humans and rodents, the ramifications of excess nickel in the environment for animal and human health remain largely undescribed. Nickel and other cationic metals travel through waterways and bind to soils and sediments. To evaluate the potential toxic effects of nickel at environmental contaminant levels (8.9-7,600 µg Ni/g dry weight of sediment and 50-800 µg NiCl2/L of water), we conducted assays using two cosmopolitan nematodes, Caenorhabditis elegans and Pristionchus pacificus. We assayed the effects of both sediment-bound and aqueous nickel upon animal growth, developmental survival, lifespan, and fecundity. Uncontaminated sediments were collected from sites in the Midwestern United States and spiked with a range of nickel concentrations. We found that nickel-spiked sediment substantially impairs both survival from larval to adult stages and adult longevity in a concentration-dependent manner. Further, while aqueous nickel showed no adverse effects on either survivorship or longevity, we observed a significant decrease in fecundity, indicating that aqueous nickel could have a negative impact on nematode physiology. Intriguingly, C. elegansand P. pacificus exhibit similar, but not identical, responses to nickel exposure. Moreover, P. pacificus could be tested successfully in sediments inhospitable to C. elegans. Our results add to a growing body of literature documenting the impact of nickel on animal physiology, and suggest that environmental toxicological studies could gain an advantage by widening their repertoire of nematode species.

  9. Two light, sweet Indonesian crudes assayed

    SciTech Connect

    Rhodes, A.K.

    1994-10-31

    Two crudes from Southeast Asia have been assayed. Belida, from the South Natuna Sea, is a light, sweet crude, with an API gravity of 45.1 [degree] and almost no sulfur (0.02 wt %). Hydra, from the Zone of Cooperation between Indonesia and Australia in the Timor Sea, has a gravity of 37.5 API and a sulfur content of 0.08 wt%. Belida operator Conoco Indonesia Ltd. began phase-two oil flow from a directional development well last October. Production from the field was 84,665 b/d on Oct. 21, 1993, and was expected to reach 90,000 b/d by the end of that month. The operator expected a peak of 100,00 b/d some time this year. The second crude, Hydra, is from the first well drilled in the Zone of Cooperation between Indonesia and Australia. Marathon Oil Co. spudded the first Hydra well in block ZOCA 9-11 in late 1992. As of early last year, five operating groups were expected to drill seven wells in second-half 1993. And a total of 26 wells has been committed for the Zone of Cooperation between 1995 and 1997, making the area a hotbed of exploration. Although the Journal has acquired no additional information on the Hydra program since that time, the assay may provided an idea of the quality of the crudes from that area.

  10. Assay optimization: a statistical design of experiments approach.

    PubMed

    Altekar, Maneesha; Homon, Carol A; Kashem, Mohammed A; Mason, Steven W; Nelson, Richard M; Patnaude, Lori A; Yingling, Jeffrey; Taylor, Paul B

    2007-03-01

    With the transition from manual to robotic HTS in the last several years, assay optimization has become a significant bottleneck. Recent advances in robotic liquid handling have made it feasible to reduce assay optimization timelines with the application of statistically designed experiments. When implemented, they can efficiently optimize assays by rapidly identifying significant factors, complex interactions, and nonlinear responses. This article focuses on the use of statistically designed experiments in assay optimization.

  11. Performance Characteristics of Xpert Flu/RSV XC Assay.

    PubMed

    Popowitch, Elena B; Miller, Melissa B

    2015-08-01

    The Xpert Flu/RSV XC assay was compared to laboratory-developed tests (LDTs) (n = 207) and the Xpert Flu assay (n = 147) using archived nasopharyngeal swabs. The percentages of positive agreements with LDTs were 97.8% for influenza A, 97.2% for influenza B, and 89.3% for RSV. The sensitivity of influenza detection was improved with the Xpert Flu/RSV XC assay compared to the Xpert Flu assay.

  12. Development of an immunofluorescence focus assay for Ebola virus.

    PubMed Central

    Truant, A L; Regnery, R L; Kiley, M P

    1983-01-01

    A 48-h indirect immunofluorescence focus assay for the quantitation of Ebola virus was developed, utilizing HeLa-229 cell monolayers. The dose dependency and the sensitivity of this assay as compared with conventional assays are reported. This indirect immunofluorescence focus assay can be used as a rapid, quantitative test for the detection of Ebola virus, an agent from Africa known to cause hemorrhagic fever. Images PMID:6352735

  13. Hormone assays: some aspects that endocrinologists should know.

    PubMed

    Alfayate, Rocío; Mauri, Montserrat

    2008-02-01

    Since the pioneering works of Yalow and Berson that introduced radioimmunoassays (RIA), hormone assays have been developed gradually, with improvements in all aspects of their design, from immunoradiometric assays to automatization. Examples of this evolution are the thyrotropin (TSH) and parathyroid (PTH) assays. Despite the strong accuracy and reliability of currently used hormone assays, some limitations should be reviewed, such as interference by autoantibodies, heterophile antibodies or macroprolactin and the hook effect.

  14. MAGNETIC DENSITOMETER

    DOEpatents

    McCann, J.A.; Jones, R.H.

    1961-08-15

    A magnetic densitometer for locating defects and metallic inclusions in materials is described. The apparatus consists of two primary coils connected in series opposition and adapted te be placed in inductive relation to the material under test, a source of constant frequency alternating current coupled across the primary coil combination, a pick-up coil disposed in symmetrical inductive relationship with said primary coils, a phase-shifter coupled to the output of the energizing source. The output of the phase-shifter is coupled in series with the pick-up coil. An amplifier is provided selective to the third harmonic of the energizing source frequency. The series combination of the pick-up coil and the phase-shifter output are connected across the input of the amplifier, and an amplitude comparitor is coupled to the output of the amplifier and the energizing source for comparing the instantaneous amplitude of the amplifier output and the instantaneous output of the energizing source and producing an output proportional to the difference in amplitude. A recorder is coupled to the output of the amplitude comparison means to give an indication of the amplitude difference, thereby providing a permanent presentation of the character of the changes in characteristics exhibited by the material under test. (AEC)

  15. Comparison of two rapid assays for Clostridium difficile Common antigen and a C difficile toxin A/B assay with the cell culture neutralization assay.

    PubMed

    Reller, Megan E; Alcabasa, Romina C; Lema, Clara A; Carroll, Karen C

    2010-01-01

    We compared 3 rapid assays for Clostridium difficile with a cell culture cytotoxicity neutralization assay (CCNA). Of 600 stool samples, 46 were positive for toxigenic C difficile. Both rapid common antigen assays were highly sensitive (91.3%-100%) and, therefore, were appropriate screening tests. The rapid toxin assay had poor sensitivity (61%) but excellent specificity (99.3%). Testing stools for glutamate dehydrogenase (step 1) and those positive with a rapid toxin assay (step 2) would correctly classify 81% of submitted specimens within 2 hours, including during periods of limited staffing (evenings, nights, and weekends). CCNA could then be used as a third step to test rapid toxin-negative samples, thereby providing a final result for the remaining 19% of samples by 48 to 72 hours. The use of rapid assays as outlined could enhance timely diagnosis of C difficile.

  16. 21 CFR 866.2350 - Microbiological assay culture medium.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... organism in the innoculated medium. Test results aid in the diagnosis of disease resulting from...

  17. 21 CFR 866.2350 - Microbiological assay culture medium.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... organism in the innoculated medium. Test results aid in the diagnosis of disease resulting from...

  18. 21 CFR 866.2350 - Microbiological assay culture medium.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... organism in the innoculated medium. Test results aid in the diagnosis of disease resulting from...

  19. 21 CFR 866.2350 - Microbiological assay culture medium.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... organism in the innoculated medium. Test results aid in the diagnosis of disease resulting from...

  20. 21 CFR 866.2350 - Microbiological assay culture medium.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Microbiological assay culture medium. 866.2350... Microbiological assay culture medium. (a) Identification. A microbiological assay culture medium is a device that... organism in the innoculated medium. Test results aid in the diagnosis of disease resulting from...

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

    PubMed

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

    2009-06-01

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

  2. Assay for mutagenesis in heterozygous diploid human lymphoblasts

    DOEpatents

    Skopek, Thomas R.; Liber, Howard L.; Penman, Bruce W.; Thilly, William G.; Hoppe, IV, Henry

    1981-01-01

    An assay is disclosed for determining mutagenic damage caused by the administration of a known or suspected mutagen to diploid human lymphoblastoid cell lines. The gene locus employed for this assay is the gene for thymidine kinase, uridine kinase, or cytidine deaminase. Since human lymphoblastoid cells contain two genes for these enzymes, heterozygotes of human lymphoblastoid cells are used in this assay.

  3. 21 CFR 864.7500 - Whole blood hemoglobin assays.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Whole blood hemoglobin assays. 864.7500 Section... blood hemoglobin assays. (a) Identification. A whole blood hemoglobin assay is a device consisting or... hemoglobin content of whole blood for the detection of anemia. This generic device category does not...

  4. 21 CFR 864.7500 - Whole blood hemoglobin assays.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Whole blood hemoglobin assays. 864.7500 Section... blood hemoglobin assays. (a) Identification. A whole blood hemoglobin assay is a device consisting or... hemoglobin content of whole blood for the detection of anemia. This generic device category does not...

  5. 21 CFR 864.7500 - Whole blood hemoglobin assays.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Whole blood hemoglobin assays. 864.7500 Section... blood hemoglobin assays. (a) Identification. A whole blood hemoglobin assay is a device consisting or... hemoglobin content of whole blood for the detection of anemia. This generic device category does not...

  6. 21 CFR 864.7500 - Whole blood hemoglobin assays.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Whole blood hemoglobin assays. 864.7500 Section... blood hemoglobin assays. (a) Identification. A whole blood hemoglobin assay is a device consisting or... hemoglobin content of whole blood for the detection of anemia. This generic device category does not...

  7. 21 CFR 864.7500 - Whole blood hemoglobin assays.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Whole blood hemoglobin assays. 864.7500 Section... blood hemoglobin assays. (a) Identification. A whole blood hemoglobin assay is a device consisting or... hemoglobin content of whole blood for the detection of anemia. This generic device category does not...

  8. 21 CFR 864.7100 - Red blood cell enzyme assay.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Red blood cell enzyme assay. 864.7100 Section 864...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell enzyme assay. (a) Identification. Red blood cell enzyme assay is a device used to measure the activity...

  9. 21 CFR 864.7100 - Red blood cell enzyme assay.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Red blood cell enzyme assay. 864.7100 Section 864...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell enzyme assay. (a) Identification. Red blood cell enzyme assay is a device used to measure the activity...

  10. 21 CFR 864.7100 - Red blood cell enzyme assay.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Red blood cell enzyme assay. 864.7100 Section 864...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell enzyme assay. (a) Identification. Red blood cell enzyme assay is a device used to measure the activity...

  11. 21 CFR 864.7100 - Red blood cell enzyme assay.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Red blood cell enzyme assay. 864.7100 Section 864...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell enzyme assay. (a) Identification. Red blood cell enzyme assay is a device used to measure the activity...

  12. 21 CFR 864.7100 - Red blood cell enzyme assay.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Red blood cell enzyme assay. 864.7100 Section 864...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7100 Red blood cell enzyme assay. (a) Identification. Red blood cell enzyme assay is a device used to measure the activity...

  13. A Different Approach to Validating Screening Assays for Developmental Toxicity

    EPA Science Inventory

    BACKGROUND: There continues to be many efforts around the world to develop assays that are shorter than the traditional embryofetal developmental toxicity assay, or use fewer or no mammals, or use less compound, or have all three attributes. Each assay developer needs to test th...

  14. Systems, devices, and methods for agglutination assays using sedimentation

    DOEpatents

    Schaff, Ulrich Y.; Sommer, Gregory J.; Singh, Anup K.

    2016-01-26

    Embodiments of the present invention include methods for conducting agglutination assays using sedimentation. Aggregates may be exposed to sedimentation forces and travel through a density medium to a detection area. Microfluidic devices, such as microfluidic disks, are described for conducting the agglutination assays, as are systems for conducting the assays.

  15. Antibody secreting cell assay for influenza A virus in swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An ELISPOT assay to enumerate B-cells producing antibodies specific to a given antigen, also known as an antibody secreting cell (ASC) assay, was adapted to detect B-cells specific for influenza A virus (IAV). The assay is performed ex vivo and enumerates ASC at a single cell level. A simple ASC det...

  16. Label-Free Isothermal Amplification Assay for Specific and Highly Sensitive Colorimetric miRNA Detection

    PubMed Central

    2016-01-01

    We describe a new method for the detection of miRNA in biological samples. This technology is based on the isothermal nicking enzyme amplification reaction and subsequent hybridization of the amplification product with gold nanoparticles and magnetic microparticles (barcode system) to achieve naked-eye colorimetric detection. This platform was used to detect a specific miRNA (miRNA-10b) associated with breast cancer, and attomolar sensitivity was demonstrated. The assay was validated in cell culture lysates from breast cancer cells and in serum from a mouse model of breast cancer. PMID:27713932

  17. Magnetic-flux pump

    NASA Technical Reports Server (NTRS)

    Hildebrandt, A. F.; Elleman, D. D.; Whitmore, F. C. (Inventor)

    1966-01-01

    A magnetic flux pump is described for increasing the intensity of a magnetic field by transferring flux from one location to the magnetic field. The device includes a pair of communicating cavities formed in a block of superconducting material, and a piston for displacing the trapped magnetic flux into the secondary cavity producing a field having an intense flux density.

  18. Magnetic multilayer structure

    DOEpatents

    Herget, Philipp; O'Sullivan, Eugene J.; Romankiw, Lubomyr T.; Wang, Naigang; Webb, Bucknell C.

    2016-07-05

    A mechanism is provided for an integrated laminated magnetic device. A substrate and a multilayer stack structure form the device. The multilayer stack structure includes alternating magnetic layers and diode structures formed on the substrate. Each magnetic layer in the multilayer stack structure is separated from another magnetic layer in the multilayer stack structure by a diode structure.

  19. Superconducting magnets 1992

    SciTech Connect

    Not Available

    1993-06-01

    This report discusses the following topics on Superconducting Magnets; SSC Magnet Industrialization; Collider Quadrupole Development; A Record-Setting Magnet; D20: The Push Beyond 10T; Nonaccelerator Applications; APC Materials Development; High-T{sub c} at Low Temperature; Cable and Cabling-Machine Development; and Analytical Magnet Design.

  20. Contactless Magnetic Slip Ring

    NASA Technical Reports Server (NTRS)

    Kumagai, Hiroyuki (Inventor); Deardon, Joe D. (Inventor)

    1997-01-01

    A contactless magnetic slip ring is disclosed having a primary coil and a secondary coil. The primary and secondary coils are preferably magnetically coupled together, in a highly reliable efficient manner, by a magnetic layered core. One of the secondary and primary coils is rotatable and the contactless magnetic slip ring provides a substantially constant output.

  1. A Magnetic Paradox

    ERIC Educational Resources Information Center

    Arndt, Ebe

    2006-01-01

    Two recent articles in this journal described how an air core solenoid connected to an ac power source may restore the magnetization of a bar magnet with an alternating magnetic field (see Figs. 1 and 2). Although we are quite accustomed to using a constant magnetic field in an air core solenoid to remagnetize a ferromagnet, it is puzzling that we…

  2. Human tear analysis with miniaturized multiplex cytokine assay on “wall-less” 96-well plate

    PubMed Central

    Quah, Joanne; Tong, Louis; Kim, Namyong

    2015-01-01

    Purpose Tears are a particularly limited body fluid and commonly used in the diagnosis of patients who have ocular diseases. A popular method for analysis of ocular inflammation in tears uses Luminex® bead multiplex technology to generate valuable multiple cytokine profile outputs with 25–50 µl tear sample volume. We propose a method for measuring tear cytokines with 5 μl tear sample volume and 80% reduced Luminex reagents compared to previous protocols. Methods Using human tears pooled from 1,000 participants, the DA-Bead-based method running at 5–20 µl volume, using manual pipetting, in conjunction with a magnetic Luminex cytokine (four-plex) panel assay in a 96-well format was performed and validated for tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-1β, and IL-6. Results Upon use of the DA-Bead method at the 5 μl volume with cytokine standards, the concentrations of each of the four cytokines were found to be linear over a range of 3.5–4 log pg/ml with an intra-assay coefficient of variation (CV) ≤5%, inter-assay %CV ≤10%, and accuracy within the 70–130% range. Upon use of a 5 µl healthy pooled tear sample, cytokine concentrations were detected with a precision intra-assay %CV ˂ 20% for IL-6, IFN-γ, or TNF-α or 30.37% with IL-1β. The inter-assay %CV with tears was ≤20.84% for all cytokines. Tear volumes run at 5 μl on DA-Bead produced a similar cytokine expression profile at a 1-month interval and were highly correlated with the larger 10 μl–based tear sample volume cytokine profile with R2 = 0.98. Conclusions DA-Bead assay is highly sensitive and reproducible and has a performance profile that is potentially suitable for use in standard clinical scenarios. Considering the use of as little as 5 µl of assay beads and 5 µl sample, this is also likely to reduce the assay cost significantly and ease diagnosis of patients with ocular diseases. PMID:26539027

  3. Scanning magnetic tunnel junction microscope for high-resolution imaging of remanent magnetization fields

    NASA Astrophysics Data System (ADS)

    Lima, E. A.; Bruno, A. C.; Carvalho, H. R.; Weiss, B. P.

    2014-10-01

    Scanning magnetic microscopy is a new methodology for mapping magnetic fields with high spatial resolution and field sensitivity. An important goal has been to develop high-performance instruments that do not require cryogenic technology due to its high cost, complexity, and limitation on sensor-to-sample distance. Here we report the development of a low-cost scanning magnetic microscope based on commercial room-temperature magnetic tunnel junction (MTJ) sensors that typically achieves spatial resolution better than 7 µm. By comparing different bias and detection schemes, optimal performance was obtained when biasing the MTJ sensor with a modulated current at 1.0 kHz in a Wheatstone bridge configuration while using a lock-in amplifier in conjunction with a low-noise custom-made preamplifier. A precision horizontal (x-y) scanning stage comprising two coupled nanopositioners controls the position of the sample and a linear actuator adjusts the sensor-to-sample distance. We obtained magnetic field sensitivities better than 150 nT/Hz1/2 between 0.1 and 10 Hz, which is a critical frequency range for scanning magnetic microscopy. This corresponds to a magnetic moment sensitivity of 10-14 A m2, a factor of 100 better than achievable with typical commercial superconducting moment magnetometers. It also represents an improvement in sensitivity by a factor between 10 and 30 compared to similar scanning MTJ microscopes based on conventional bias-detection schemes. To demonstrate the capabilities of the instrument, two polished thin sections of representative geological samples were scanned along with a synthetic sample containing magnetic microparticles. The instrument is usable for a diversity of applications that require mapping of samples at room temperature to preserve magnetic properties or viability, including paleomagnetism and rock magnetism, nondestructive evaluation of materials, and biological assays.

  4. Linear magnetic bearing

    NASA Technical Reports Server (NTRS)

    Studer, P. A. (Inventor)

    1983-01-01

    A linear magnetic bearing system having electromagnetic vernier flux paths in shunt relation with permanent magnets, so that the vernier flux does not traverse the permanent magnet, is described. Novelty is believed to reside in providing a linear magnetic bearing having electromagnetic flux paths that bypass high reluctance permanent magnets. Particular novelty is believed to reside in providing a linear magnetic bearing with a pair of axially spaced elements having electromagnets for establishing vernier x and y axis control. The magnetic bearing system has possible use in connection with a long life reciprocating cryogenic refrigerator that may be used on the space shuttle.

  5. Ovariectomized mouse uterotrophic assay of 36 chemicals.

    PubMed

    Ohta, Ryo; Takagi, Atsuya; Ohmukai, Hideo; Marumo, Hideki; Ono, Atsushi; Matsushima, Yuko; Inoue, Tohru; Ono, Hiroshi; Kanno, Jun

    2012-01-01

    The concern over endocrine disruptors prompted international establishment of a strategic framework for the identification of the estrogenic compounds. OECD has launched the Conceptual Framework tool box containing various screening and testing methods including the uterotrophic assay. The (anti)estrogenicity of 36 chemicals suspected to be estrogen-receptor interactive by in silico and/or in vitro screening in the Extended Scheme for Endocrine Disruptor Screening and Testing of the Ministry of Health, Labour and Welfare, Japan, were monitored by the uterotrophic assay using C57BL/6J ovariectomized adult female mice after a 7-day exposure by oral gavage (po) and subcutaneous injection (sc). Ethynyl estradiol was used as reference for agonist and antagonist detection. In addition, Bisphenol A (sc) and Genistein (po) were tested for the comparison to rat assays. Among the 36, 2-[Bis(4-hydroxy-phenyl)methyl]benzylalcohol, 2,2',4,4'-Tetrahydroxybenzophenone, 2,4-Dihydroxybenzophenone, 3,3',5-Triiodothyroacetic acid, New fuchsin and alpha-Naphtholbenzein, showed both estrogenic agonistic and antagonistic activities; first two showed U-shaped dose-response in antagonistic studies. N,N-Diphenyl-p-phenylenediamine, 2,2'-Dihydroxy-4,4'-dimethoxybenzophenone, n-Butyl 4-hydroxybenzoate, and Reserpine were agonistic by sc. Benzo [a] pyrene, Benz [a] anthracene, Dibenz [a,h] anthracene, 2-(2H-Benzotriazol-2-yl)-4,6-di(t-pentyl)phenol, Rosemarinic acid, meta-Thymol, 6-Gingerol, Colchicine, Malachite green base, Fenbuconazole, and Lead acetate were antagonistic. The rest, i.e. n-Heptyl 4-hydroxybenzoate, Tetrazolium violet, Pravastatin sodium salt, Physostigmine, salicylate (1:1), Nordihydroguaiaretic acid, o-Cresolphthalein, 1,3-Dinitrobenzene, C.I. Pigment orange, Tetrabromobis-phenol-A, 2-Hydroxy-4-methoxybenzophenone, Ethylparaben, Propyl p-hydroxybenzoate, Kaempferol, 2-(2-Benzotriazolyl)-p-cresol and Phenolphthalein were negative for both effects. Taking together with in

  6. Hybrid superconducting magnetic suspensions

    SciTech Connect

    Tixador, P.; Hiebel, P.; Brunet, Y.

    1996-07-01

    Superconductors, especially high T{sub c} ones, are the most attractive materials to design stable and fully passive magnetic suspensions which have to control five degrees of freedom. The hybrid superconducting magnetic suspensions present high performances and a simple cooling mode. They consist of a permanent magnet bearing, stabilized by a suitable magnet-superconductor structure. Several designs are given and compared in terms of forces and stiffnesses. The design of the magnet bearing plays an important part. The superconducting magnetic bearing participates less in levitation but must provide a high stabilizing stiffness. This is achieved by the magnet configuration, a good material in term of critical current density and field cooling. A hybrid superconducting suspension for a flywheel is presented. This system consists of a magnet thrust bearing stabilized by superconductors interacting with an alternating polarity magnet structure. First tests and results are reported. Superconducting materials are magnetically melt-textured YBaCuO.

  7. Engineering biofunctional magnetic nanoparticles for biotechnological applications

    NASA Astrophysics Data System (ADS)

    Moros, Maria; Pelaz, Beatriz; López-Larrubia, Pilar; García-Martin, Maria L.; Grazú, Valeria; de La Fuente, Jesus M.

    2010-09-01

    Synthesis and characterization of magnetic nanoparticles with excellent size control are showed here. Their functionalization using an amphiphilic polymer is also described. This strategy allows the stabilization of magnetic nanoparticles in aqueous solvents and in addition, the polymer shell serves as a platform to incorporate relevant biomolecules, such as poly(ethylene glycol) and a number of carbohydrates. Nanoparticles functionalized with carbohydrates show the ability to avoid unspecific interactions between proteins present in the working medium and the nanoparticles, so can be used as an alternative to poly(ethylene glycol) molecules. Results confirm these nanoparticles as excellent contrast agents for magnetic resonance imaging. Changes in the spin-spin transversal relaxation times of the surrounding water protons due to nanoparticle aggregation demonstrates the bioactivity of these nanoparticles functionalized with carbohydrates. To finish with, nanoparticle toxicity is evaluated by means of MTT assay. The obtained results clearly indicate that these nanoparticles are excellent candidates for their further application in nanomedicine or nanobiotechnology.Synthesis and characterization of magnetic nanoparticles with excellent size control are showed here. Their functionalization using an amphiphilic polymer is also described. This strategy allows the stabilization of magnetic nanoparticles in aqueous solvents and in addition, the polymer shell serves as a platform to incorporate relevant biomolecules, such as poly(ethylene glycol) and a number of carbohydrates. Nanoparticles functionalized with carbohydrates show the ability to avoid unspecific interactions between proteins present in the working medium and the nanoparticles, so can be used as an alternative to poly(ethylene glycol) molecules. Results confirm these nanoparticles as excellent contrast agents for magnetic resonance imaging. Changes in the spin-spin transversal relaxation times of the

  8. On-chip determination of C-reactive protein using magnetic particles in continuous flow.

    PubMed

    Phurimsak, Chayakom; Tarn, Mark D; Peyman, Sally A; Greenman, John; Pamme, Nicole

    2014-11-01

    We demonstrate the application of a multilaminar flow platform, in which functionalized magnetic particles are deflected through alternating laminar flow streams of reagents and washing solutions via an external magnet, for the rapid detection of the inflammatory biomarker, C-reactive protein (CRP). The two-step sandwich immunoassay was accomplished in less than 60 s, a vast improvement on the 80-300 min time frame required for enzyme-linked immunosorbent assays (ELISA) and the 50 min necessary for off-chip magnetic particle-based assays. The combination of continuous flow and a stationary magnet enables a degree of autonomy in the system, while a detection limit of 0.87 μg mL(-1) makes it suitable for the determination of CRP concentrations in clinical diagnostics. Its applicability was further proven by assaying real human serum samples and comparing those results to values obtained using standard ELISA tests.

  9. High efficiency magnetic bearings

    NASA Technical Reports Server (NTRS)

    Studer, Philip A.; Jayaraman, Chaitanya P.; Anand, Davinder K.; Kirk, James A.

    1993-01-01

    Research activities concerning high efficiency permanent magnet plus electromagnet (PM/EM) pancake magnetic bearings at the University of Maryland are reported. A description of the construction and working of the magnetic bearing is provided. Next, parameters needed to describe the bearing are explained. Then, methods developed for the design and testing of magnetic bearings are summarized. Finally, a new magnetic bearing which allows active torque control in the off axes directions is discussed.

  10. Stacked magnet superconducting bearing

    SciTech Connect

    Rigney, T.K. II; Saville, M.P.

    1993-06-15

    A superconducting bearing is described, comprising: a plurality of permanent magnets magnetized end-to-end and stacked side-by-side in alternating polarity, such that flux lines flow between ends of adjacent magnets; isolating means, disposed between said adjacent magnets, for reducing flux leakage between opposing sides of said adjacent magnets; and a member made of superconducting material having at least one surface in communication with said flux lines.

  11. Magnetic Damping For Maglev

    DOE PAGES

    Zhu, S.; Cai, Y.; Rote, D. M.; Chen, S. S.

    1998-01-01

    Magnetic damping is one of the important parameters that control the response and stability of maglev systems. An experimental study to measure magnetic damping directly is presented. A plate attached to a permanent magnet levitated on a rotating drum was tested to investigate the effect of various parameters, such as conductivity, gap, excitation frequency, and oscillation amplitude, on magnetic damping. The experimental technique is capable of measuring all of the magnetic damping coefficients, some of which cannot be measured indirectly.

  12. Facility Measures Magnetic Fields

    NASA Technical Reports Server (NTRS)

    Honess, Shawn B.; Narvaez, Pablo; Mcauley, James M.

    1991-01-01

    Partly automated facility measures and computes steady near magnetic field produced by object. Designed to determine magnetic fields of equipment to be installed on spacecraft including sensitive magnetometers, with view toward application of compensating fields to reduce interfernece with spacecraft-magnetometer readings. Because of its convenient operating features and sensitivity of its measurements, facility serves as prototype for similar facilities devoted to magnetic characterization of medical equipment, magnets for high-energy particle accelerators, and magnetic materials.

  13. Magnetically operated check valve

    NASA Technical Reports Server (NTRS)

    Morris, Brian G. (Inventor); Bozeman, Richard J., Jr. (Inventor)

    1994-01-01

    A magnetically operated check valve is disclosed. The valve is comprised of a valve body and a movable poppet disposed therein. A magnet attracts the poppet to hold the valve shut until the force of fluid flow through the valve overcomes the magnetic attraction and moves the poppet to an unseated, open position. The poppet and magnet are configured and disposed to trap a magnetically attracted particulate and prevent it from flowing to a valve seating region.

  14. Accuracy of the endpoint assay for virus titration.

    PubMed

    Nielsen, L K; Smyth, G K; Greenfield, P F

    1992-01-01

    The statistics of estimators used with the endpoint assay for virus titration were investigated. For a standard assay with 10 wells/dilution, the graphical estimator traditionally used was found to produce estimates with significant positive bias and a relatively low accuracy. Furthermore, the graphical estimator was found to be inconsistent. A superior estimator based on the maximum likelihood principle was developed. The results are discussed in relation to the choice between the endpoint titration assay and the plaque assay, and an alternative two-stage assay is presented.

  15. Antioxidant assay using genetically engineered bioluminescent Escherichia coli

    NASA Astrophysics Data System (ADS)

    Bartolome, Amelita; Macalino, Bernadette; Pastoral, Ian Lemuel; Sevilla, Fortunato, III

    2006-02-01

    A new antioxidant activity assay based on the reactive oxygen species (ROS)-inducible bacterial strain (E. coli DPD2511) is described. The strain harbors the plasmid pKatG::luxCDABE and responds to hydrogen peroxide treatment by increasing light emission at 490 nm. Antioxidant capacity is evaluated through the ability of an agent to inhibit the hydrogen peroxide-induced bioluminescence of E. coli DPD2511. Applicability of the developed assay in detecting levels of antioxidants in various aqueous plant extracts is demonstrated. The assay was validated against 2,2-diphenylpicrylhydrazyl (DPPH) assay, a known antioxidant assay.

  16. Universal fieldable assay with unassisted visual detection

    NASA Technical Reports Server (NTRS)

    Chelyapov, Nicolas (Inventor)

    2012-01-01

    A universal detection system based on allosteric aptamers, signal amplification cascade, and eye-detectable phrase transition. A broadly applicable homogeneous detection system is provided. It utilizes components of the blood coagulation cascade in the presence of polystyrene microspheres (MS) as a signal amplifier. Russell's viper venom factor X activator (RVV-X) triggers the cascade, which results in an eye-visible phase transition--precipitation of MS bound to clotted fibrin. An allosteric RNA aptamer, RNA132, with affinity for RVV-X and human vascular endothelial growth factor (VEGF.sub.165) was created. RNA132 inhibits enzymatic activity of RVV-X. The effector molecule, VEGF.sub.165, reverses the inhibitory activity of RNA132 on RVV-X and restores its enzymatic activity, thus triggering the cascade and enabling the phase transition. Similar results were obtained for another allosteric aptamer modulated by a protein tyrosine phosphatase. The assay is instrumentation-free for both processing and readout.

  17. Rapid Automated Sample Preparation for Biological Assays

    SciTech Connect

    Shusteff, M

    2011-03-04

    Our technology utilizes acoustic, thermal, and electric fields to separate out contaminants such as debris or pollen from environmental samples, lyse open cells, and extract the DNA from the lysate. The objective of the project is to optimize the system described for a forensic sample, and demonstrate its performance for integration with downstream assay platforms (e.g. MIT-LL's ANDE). We intend to increase the quantity of DNA recovered from the sample beyond the current {approx}80% achieved using solid phase extraction methods. Task 1: Develop and test an acoustic filter for cell extraction. Task 2: Develop and test lysis chip. Task 3: Develop and test DNA extraction chip. All chips have been fabricated based on the designs laid out in last month's report.

  18. Nondestructive Assay Options for Spent Fuel Encapsulation

    SciTech Connect

    Tobin, Stephen J.; Jansson, Peter

    2014-10-02

    This report describes the role that nondestructive assay (NDA) techniques and systems of NDA techniques may have in the context of an encapsulation and deep geological repository. The potential NDA needs of an encapsulation and repository facility include safeguards, heat content, and criticality. Some discussion of the facility needs is given, with the majority of the report concentrating on the capability and characteristics of individual NDA instruments and techniques currently available or under development. Particular emphasis is given to how the NDA techniques can be used to determine the heat production of an assembly, as well as meet the dual safeguards needs of 1) determining the declared parameters of initial enrichment, burn-up, and cooling time and 2) detecting defects (total, partial, and bias). The report concludes with the recommendation of three integrated systems that might meet the combined NDA needs of the encapsulation/repository facility.

  19. A miniaturized fibrinolytic assay for plasminogen activators

    NASA Technical Reports Server (NTRS)

    Lewis, M. L.; Nachtwey, D. S.; Damron, K. L.

    1991-01-01

    This report describes a micro-clot lysis assay (MCLA) for evaluating fibrinolytic activity of plasminogen activators (PA). Fibrin clots were formed in wells of microtiter plates. Lysis of the clots by PA, indicated by change in turbidity (optical density, OD), was monitored with a microplate reader at five minutes intervals. Log-log plots of PA dilution versus endpoint, the time at which the OD value was halfway between the maximum and minimum value for each well, were linear over a broad range of PA concentrations (2-200 International units/ml). The MCLA is a modification and miniaturization of well established fibrinolytic methods. The significant practical advantages of the MCLA are that it is a simple, relatively sensitive, non-radioactive, quantitative, kinetic, fibrinolytic micro-technique which can be automated.

  20. Analyte detection using an active assay

    DOEpatents

    Morozov, Victor; Bailey, Charles L.; Evanskey, Melissa R.

    2010-11-02

    Analytes using an active assay may be detected by introducing an analyte solution containing a plurality of analytes to a lacquered membrane. The lacquered membrane may be a membrane having at least one surface treated with a layer of polymers. The lacquered membrane may be semi-permeable to nonanalytes. The layer of polymers may include cross-linked polymers. A plurality of probe molecules may be arrayed and immobilized on the lacquered membrane. An external force may be applied to the analyte solution to move the analytes towards the lacquered membrane. Movement may cause some or all of the analytes to bind to the lacquered membrane. In cases where probe molecules are presented, some or all of the analytes may bind to probe molecules. The direction of the external force may be reversed to remove unbound or weakly bound analytes. Bound analytes may be detected using known detection types.