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Sample records for double antibody method

  1. Chromosome doubling method

    DOEpatents

    Kato, Akio

    2006-11-14

    The invention provides methods for chromosome doubling in plants. The technique overcomes the low yields of doubled progeny associated with the use of prior techniques for doubling chromosomes in plants such as grasses. The technique can be used in large scale applications and has been demonstrated to be highly effective in maize. Following treatment in accordance with the invention, plants remain amenable to self fertilization, thereby allowing the efficient isolation of doubled progeny plants.

  2. Double-antigen sandwich ELISA for the detection of anti-hepatitis C virus antibodies.

    PubMed

    He, Jing; Xiu, Bingshui; Wang, Guohua; Chen, Kun; Feng, Xiaoyan; Song, Xiaoguo; Zhu, Cuixia; Ling, Shigan; Zhang, Heqiu

    2011-01-01

    A double-antigen sandwich ELISA was developed a detection of HCV antibodies by a recombinant multi-epitope HCV antigen and a biotin-streptavidin amplification system. Three plasma specimens from 1708 individuals who were suspected previously to be HCV-positive using an HCV antibody diagnostic kit (Chuangxin, Xiamen, China) displayed negative results when using the ELISA. These results were validated by a recombinant immunoblotting assay (two were negative, and one was indeterminate). Among 889 blood specimens donated for clinical evaluation, 246 were positive and 630 were negative using the ELISA. The sensitivity and specificity of the ELISA were 98.7% and 100%, respectively. In 43 donors and 14 patients with chronic hepatitis C, the detectable rates for HCV IgM by both ELISA and the HCV anti-IgM detection reagents (Huimin, Shenyang, China) were 100%, and the detectable rate for HCV IgG using an indirect HCV-antibody detection kit (GWK, Beijing, China) was 98.3%. Thus, the double-antigen sandwich ELISA exhibits strong specificity and sensitivity and has been approved by the China State Food and Drug Administration (SFDA). The performance of the double-antigen sandwich ELISA was similar to the Ortho ELISA 3.0. It did not give false-negative results otherwise IgM was undetectable using an indirect HCV-antibody detection kit. This ELISA provides another method for the detection of HCV antibodies. PMID:21029749

  3. Methods of identification employing antibody profiles

    DOEpatents

    Francoeur, Ann-Michele

    1993-12-14

    An identification method, applicable to the identification of animals or inanimate objects, is described. The method takes advantage of the set of individual-specific antibodies that are part of the unique antibody repertoire present in animals, by reacting an effective amount of such antibodies with a particular panel, of n-dimensional array (where n is typically one or two) consisting of an effective amount of many different antigens (typically greater than one thousand), to give antibody-antigen complexes. The profile or pattern formed by the antigen-antibody complexes, termed an antibody fingerprint, when revealed by an effective amount of an appropriate detector molecule, is uniquely representative of a particular individual. The method can similarly be used to distinguish genetically, or otherwise similar individuals, or their body parts containing individual-specific antibodies.

  4. Individual-specific antibody identification methods

    DOEpatents

    Francoeur, Ann -Michele

    1989-11-14

    An identification method, applicable to the identification of animals or inanimate objects, is described. The method takes advantage of a hithertofore unknown set of individual-specific, or IS antibodies, that are part of the unique antibody repertoire present in animals, by reacting an effective amount of IS antibodies with a particular panel, or n-dimensional array (where n is typically one or two) consisting of an effective amount of many different antigens (typically greater than one thousand), to give antibody-antigen complexes. The profile or pattern formed by the antigen-antibody complexes, termed an antibody fingerprint, when revealed by an effective amount of an appropriate detector molecule, is uniquely representative of a particular individual. The method can similarly by used to distinguish genetically, or otherwise similar individuals, or their body parts containing IS antibodies. Identification of inanimate objects, particularly security documents, is similarly affected by associating with the documents, an effective amount of a particular individual's IS antibodies, or conversely, a particular panel of antigens, and forming antibody-antigen complexes with a particular panel of antigens, or a particular individual's IS antibodies, respectively. One embodiment of the instant identification method, termed the blocked fingerprint assay, has applications in the area of allergy testing, autoimmune diagnostics and therapeutics, and the detection of environmental antigens such as pathogens, chemicals, and toxins.

  5. Method for preparation of single chain antibodies

    SciTech Connect

    Cheung, Nai-Kong V.; Guo, Hong-fen

    2012-04-03

    This invention provides a method for identifying cells expressing a target single chain antibody (scFv) directed against a target antigen from a collection of cells that includes cells that do not express the target scFv, comprising the step of combining the collection of cells with an anti-idiotype directed to an antibody specific for the target antigen and detecting interaction, if any, of the anti-idiotype with the cells, wherein the occurrence of an interaction identifies the cell as one which expresses the target scFv. This invention also provides a method for making a single chain antibody (scFv) directed against an antigen, wherein the selection of clones is made based upon interaction of those clones with an appropriate anti-idiotype, and heretofore inaccessible scFv so made. This invention provides the above methods or any combination thereof. Finally, this invention provides various uses of these methods.

  6. The Double Absorbing Boundary method

    NASA Astrophysics Data System (ADS)

    Hagstrom, Thomas; Givoli, Dan; Rabinovich, Daniel; Bielak, Jacobo

    2014-02-01

    A new approach is devised for solving wave problems in unbounded domains. It has common features to each of two types of existing techniques: local high-order Absorbing Boundary Conditions (ABC) and Perfectly Matched Layers (PML). However, it is different from both and enjoys relative advantages with respect to both. The new method, called the Double Absorbing Boundary (DAB) method, is based on truncating the unbounded domain to produce a finite computational domain Ω, and on applying a local high-order ABC on two parallel artificial boundaries, which are a small distance apart, and thus form a thin non-reflecting layer. Auxiliary variables are defined on the two boundaries and inside the layer bounded by them, and participate in the numerical scheme. The DAB method is first introduced in general terms, using the 2D scalar time-dependent wave equation as a model. Then it is applied to the 1D Klein-Gordon equation, using finite difference discretization in space and time, and to the 2D wave equation in a wave guide, using finite element discretization in space and dissipative time stepping. The computational aspects of the method are discussed, and numerical experiments demonstrate its performance.

  7. Development of a double-antibody sandwich ELISA for rapid detection of Bacillus Cereus in food.

    PubMed

    Zhu, Longjiao; He, Jing; Cao, Xiaohan; Huang, Kunlun; Luo, Yunbo; Xu, Wentao

    2016-01-01

    Bacillus cereus is increasingly recognized as one of the major causes of food poisoning in the industrialized world. In this paper, we describe a sensitive double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) that was developed for rapid detection of B. cereus in food to minimize the risk of contamination. The polyclonal antibody (pAb) and monoclonal antibodies (mAbs) specific to B. cereus were generated from rabbit antiserum and mouse ascites, respectively, using the octanoic acid/saturated ammonium sulfate precipitation method and protein A-sepharose columns. IgG-isotype mAbs were specially developed to undergo a novel peripheral multiple sites immunization for rapid gain of hybridomas and a subtractive screen was used to eliminate cross reactivity with closely related species such as Bacillus thuringiensis, B. subtilis, B. licheniformis and B. perfringens. The linear detection range of the method was approximately 1 × 10(4)-2.8 × 10(6) cells/mL with a detection limit (LOD) of 0.9 × 10(3) cells/mL. The assay was able to detect B. cereus when the samples were prepared in meat with various pathogens. The newly developed analytical method provides a rapid method to sensitively detect B. cereus in food specimens. PMID:26976753

  8. Development of a double-antibody sandwich ELISA for rapid detection of Bacillus Cereus in food

    PubMed Central

    Zhu, Longjiao; He, Jing; Cao, Xiaohan; Huang, Kunlun; Luo, Yunbo; Xu, Wentao

    2016-01-01

    Bacillus cereus is increasingly recognized as one of the major causes of food poisoning in the industrialized world. In this paper, we describe a sensitive double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) that was developed for rapid detection of B. cereus in food to minimize the risk of contamination. The polyclonal antibody (pAb) and monoclonal antibodies (mAbs) specific to B. cereus were generated from rabbit antiserum and mouse ascites, respectively, using the octanoic acid/saturated ammonium sulfate precipitation method and protein A-sepharose columns. IgG-isotype mAbs were specially developed to undergo a novel peripheral multiple sites immunization for rapid gain of hybridomas and a subtractive screen was used to eliminate cross reactivity with closely related species such as Bacillus thuringiensis, B. subtilis, B. licheniformis and B. perfringens. The linear detection range of the method was approximately 1 × 104–2.8 × 106 cells/mL with a detection limit (LOD) of 0.9 × 103 cells/mL. The assay was able to detect B. cereus when the samples were prepared in meat with various pathogens. The newly developed analytical method provides a rapid method to sensitively detect B. cereus in food specimens. PMID:26976753

  9. Method for altering antibody light chain interactions

    DOEpatents

    Stevens, Fred J.; Stevens, Priscilla Wilkins; Raffen, Rosemarie; Schiffer, Marianne

    2002-01-01

    A method for recombinant antibody subunit dimerization including modifying at least one codon of a nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in the interface segment of the light polypeptide variable region, the charged amino acid having a first polarity; and modifying at least one codon of the nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in an interface segment of the heavy polypeptide variable region corresponding to a position in the light polypeptide variable region, the charged amino acid having a second polarity opposite the first polarity. Nucleic acid sequences which code for novel light chain proteins, the latter of which are used in conjunction with the inventive method, are also provided.

  10. Double immunofluorescent staining of rat macrophages in formalin-fixed paraffin-embedded tissue using two monoclonal mouse antibodies.

    PubMed

    Isidro, Raymond A; Isidro, Angel A; Cruz, Myrella L; Hernandez, Siomara; Appleyard, Caroline B

    2015-12-01

    The conventional approach of double immunostaining to visualize more than one protein in tissues or cells using antibodies from two different host species is not always feasible due to limitations with antibody availability. Previously reported methodologies for performing multiple immunostains on the same tissue or cells with antibodies originating from the same species are varied in their complexity, sensitivity, and approach to prevent unwanted interactions between antibodies. In the ever-expanding field of macrophage biology, much more is known about mouse and human macrophages than their rat counterparts. The limited availability of validated and well-characterized monoclonal antibodies from different species is one factor responsible for preventing advances in rat macrophage biology. Here we describe an immunostaining method for identifying and examining rat macrophages that is sufficiently sensitive for use in formalin-fixed paraffin-embedded tissue and that uses only commercially available reagents and antibodies. This method can be used to help characterize both physiological and pathophysiological processes in rat macrophages and can be adapted for use with any two antibodies from the same species of origin as long as one of the antibodies is biotinylated. PMID:26403093

  11. Platelet antibody: review of detection methods

    SciTech Connect

    Schwartz, K.A.

    1988-10-01

    The driving force behind development of in vitro methods for platelet antibodies is identification of plasma factors causing platelet destruction. Early methods relied on measurement of platelet activation. Current methods are more specific and use a purified antibody against immunoglobulin or complement, which is usually labeled with /sup 125/I or tagged with an enzyme or fluorescein. Comparisons of quantitation of platelet-associated IgG show wide variability between different methods. The disparate results can be related both to differences in binding of secondary antibodies to immunoglobulin in solution compared to immunoglobulins attached to platelets and to the improper assumption that the binding ratio between the secondary detecting and primary antiplatelet antibody is one. Most assays can 1) identify neonatal isoimmune thrombocytopenia and posttransfusion purpura, 2) help to differentiate between immune and nonimmune thrombocytopenias, 3) help to sort out the offending drug when drug-induced thrombocytopenia is suspected, and 4) identify platelet alloantibodies and potential platelet donors via a cross match assay for refractory patients. However, the advantages of quantitative assays over qualitative methods with respect to predictions of patients clinical course and response to different treatments remain to be investigated. 61 references.

  12. Hidden anti-double stranded DNA antibodies in autoimmune mice.

    PubMed Central

    Fish, F; Ziff, M

    1982-01-01

    When MRL/l mouse spleen cell culture supernatants were incubated with normal mouse spleen cells, a two-50-fold increase in anti-dsDNA activity was noted. A smaller increase in anti-ssDNA and no change in anti-TNP antibody activity were observed. This 'hidden' antibody in the MRL/l supernatants could not be revealed by DNAse digestion and could not be absorbed by a DNA cellulose column. Hidden antibody was removed from supernatants by sepharose-anti Ig. After DNAase digestion of the MRL/l supernatants, hidden anti-dsDNA could not be revealed by incubation with spleen cells. All the hidden activity was excluded by gel filtration on Sephacryl S-300 (mol. wt greater than 300,000) but was banded in the low density protein area of caesium chloride equilibrium density gradients. It was also noted that MRL/l mouse sera had hidden anti-dsDNA antibodies. Hidden antibodies were present in both the IgG and IgM classes. The revealed antibodies demonstrated impaired ability to bind Fc specific anti-Ig reagents suggesting that they were partially degraded during the incubation with mouse spleen cells. The hidden anti-dsDNA thus appears to represent a DNA-anti-dsDNA complex, perhaps of very high affinity. It may explain why anti-dsDNA but not anti-ssDNA antibodies are of pathological importance in SLE. PMID:6756722

  13. Double-antibody solid-phase radioimmunoassay: a simplified phase-separation procedure applied to various ligands

    SciTech Connect

    Tevaarwerk, G.J.M.; Boyle, D.A.; Hurst, C.J.; Anguish, I.; Uksik, P.

    1980-06-01

    The purpose was to develop a simplified and reliable method of separating free from antibody-bound ligand using a precipitating antibody linked to a cellulose derivative. Dose-response curves and control sera were set up in parallel for various pituitary and placental polypeptides, steroid hormones, insulin, glucagon, triiodothyronine, thyroxine, angiotensin I, calcitonin, gastrin, cyclic AMP, and digoxin. After first-antibody reactions had reached equilibrium, free and bound ligand were separated using a double-antibody solid-phase system in parallel with conventional methods, including dextran-coated charcoal, double-antibody precipitation, single-antibody solid phase, organic solvents, salt precipitation, and anion-exchange resins. The effect of variations in temperature, incubation time, protein content, pH, and amount of separating material added were studied. The results showed that separation was complete within 1 hr for small ligand molecules and within 2 hr for larger ones. Dose-response curves and control-sera results closely paralleled those obtained with conventional methods. The method was not affected by moderate variations in incubation variables. Nonspecific binding was less than 3% in all assays, while intra-assay and interassay coefficients of variation were similar to those obtained with conventional phase-separation methods. It is concluded that the method is a simple and rapid alternative phase-separation system. It has the advantage of being free from common nonspecific intersample variations, and can be applied to any assay system based on rabbit or guinea pig antibodies without preliminay time- or reagent-consuming titration or adjustments to establish optimum phase-separating conditions.

  14. Monoclonal antibodies and method for detecting dioxins and dibenzofurans

    DOEpatents

    Vanderlaan, Martin; Stanker, Larry H.; Watkins, Bruce E.; Bailey, Nina R.

    1989-01-01

    Compositions of matter are described which include five monoclonal antibodies that react with dioxins and dibenzofurans, and the five hybridomas that produce these monoclonal antibodies. In addition, a method for the use of these antibodies in a sensitive immunoassay for dioxins and dibenzofurans is given, which permits detection of these pollutants in samples at concentrations in the range of a few parts per billion.

  15. Composition and method for detecting cancer with technetium labeled antibody fragments

    SciTech Connect

    Burchiel, S. W.; Crockford, D. R.; Rhodes, B. A.

    1984-10-23

    F(ab')/sub 2/ or Fab fragments of antibodies to: (a) human chorionic gonadotropin (hCG), hCG alpha subunit, hCG beta subunit, or an hCG-like material; or (b) other tumor specific or tumor associated molecules, to include carcinoembryonic antigen (CEA), alpha fetoprotein (AFP), human melanoma associated antigens, human sarcoma associated antigens or other antigens, are radiolabeled with technetium-99m (Tc-99m). When the F(ab')/sub 2/ or Fab fragments of antibody to such tumor associated antigens are injected intravenously into a patient, the radiolabeled composition accumulates at tumor sites. The accumulation of the cancer seeking radiopharmaceutical at tumor sites permits detection by external gamma scintigraphy. Thus, the composition is useful in the monitoring, localization and detection of cancer in the body. In an alternative composition, a double antibody approach to tumor localization using radiolabeled F(ab')/sub 2/ or Fab fragments is utilized. In this approach, a tumor specific antibody in the form of IgG, F(ab')/sub 2/ or Fab is first administered to a patient intravenously. Following a sufficient period of time, a second antibody in the form of F(ab')/sub 2/ or Fab is administered. The second antibody is radiolabeled with Tc-99m and has the property that it is reactive with the first antibody. This double antibody method has the advantage over a single antibody approach in that smaller tumors can be localized and detected and that the total amount of radioactive trace localized at the cancer site is increased.

  16. Methods to Design and Synthesize Antibody-Drug Conjugates (ADCs)

    PubMed Central

    Yao, Houzong; Jiang, Feng; Lu, Aiping; Zhang, Ge

    2016-01-01

    Antibody-drug conjugates (ADCs) have become a promising targeted therapy strategy that combines the specificity, favorable pharmacokinetics and biodistributions of antibodies with the destructive potential of highly potent drugs. One of the biggest challenges in the development of ADCs is the application of suitable linkers for conjugating drugs to antibodies. Recently, the design and synthesis of linkers are making great progress. In this review, we present the methods that are currently used to synthesize antibody-drug conjugates by using thiols, amines, alcohols, aldehydes and azides. PMID:26848651

  17. A new double views motion deblurring method

    NASA Astrophysics Data System (ADS)

    Hong, Hanyu; Hua, Xia; Zhang, Wenmo; Shi, Yu

    2015-12-01

    With improving of intelligent and automation in modern industrial production area, the detection and reconstruction of the 3D surface of the product has become an important technology, but the image which acquire on the actual production line has motion blur and this problem will affect the later reconstruction work. In order to solve this problem, a deblurring method which based on double view moving target image is proposed in this paper. We can deduce the relationship of the point spread function(PSF) path between the double view image through the epipolar geometry and the camera model. The experimental results show that deblurring with the PSF path solved by the geometric relationship achieves good results.

  18. Use of solid-phase double-antibody radioimmunoassay to identify species from small skeletal fragments.

    PubMed

    Ubelaker, Douglas H; Lowenstein, Jerold M; Hood, Darden G

    2004-09-01

    Protein radioimmunoassay (pRIA) offers the potential to identify species in small skeletal fragments submitted as forensic evidence. The technique consists of protein extraction followed by a solid-phase double-antibody radioimmunoassay using controls of antisera (raised in rabbits) and radioactive (iodine-125) antibody of rabbit gamma globulin (produced in donkeys). Species determination results from evaluation of radioactivity uptake. To demonstrate the potential of this technique, six known bone samples (three human and three nonhuman, including one from a deer [Odocoileus virginianus]) were submitted for blind analysis. pRIA correctly distinguished the human from the nonhuman samples. Using 200 mg or less of each sample, species of the deer specimen was identified correctly, given the choices of cow, deer, dog, goat, and pig. PMID:15461091

  19. Method to directly radiolabel antibodies for diagnostic imaging and therapy

    DOEpatents

    Thakur, Mathew L.

    1994-01-01

    The invention is a novel method and kit for directly radiolabeling proteins such as antibodies or antibody fragments for diagnostic and therapeutic purposes. The method comprises incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein. A kit is also provided wherein the protein and/or reducing agents may be in lyophilized form.

  20. Method to directly radiolabel antibodies for diagnostic imaging and therapy

    DOEpatents

    Thakur, Mathew L.

    1991-01-01

    The invention is a novel method and kit for directly radiolabeling proteins such as antibodies or antibody fragments for diagnostic and therapeutic purposes. The method comprises incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein. A kit is also provided wherein the protein and/or reducing agents may be in lyophilized form.

  1. Method for detection of antibodies for metallic elements

    DOEpatents

    Barrick, C.W.; Clarke, S.M.; Nordin, C.W.

    1993-11-30

    An apparatus and method for detecting antibodies specific to non-protein antigens. The apparatus is an immunological plate containing a plurality of plastic projections coated with a non-protein material. Assays utilizing the plate are capable of stabilizing the non-protein antigens with detection levels for antibodies specific to the antigens on a nanogram level. A screening assay with the apparatus allows for early detection of exposure to non-protein materials. Specifically metallic elements are detected. 10 figures.

  2. Method for detection of antibodies for metallic elements

    DOEpatents

    Barrick, Charles W.; Clarke, Sara M.; Nordin, Carl W.

    1993-11-30

    An apparatus and method for detecting antibodies specific to non-protein antigens. The apparatus is an immunological plate containing a plurality of plastic projections coated with a non-protein material. Assays utilizing the plate are capable of stabilizing the non-protein antigens with detection levels for antibodies specific to the antigens on a nanogram level. A screening assay with the apparatus allows for early detection of exposure to non-protein materials. Specifically metallic elements are detected.

  3. SPRi-based adenovirus detection using a surrogate antibody method.

    PubMed

    Abadian, Pegah N; Yildirim, Nimet; Gu, April Z; Goluch, Edgar D

    2015-12-15

    Adenovirus infection, which is a waterborne viral disease, is one of the most prevelant causes of human morbidity in the world. Thus, methods for rapid detection of this infectious virus in the environment are urgently needed for public health protection. In this study, we developed a rapid, real-time, sensitive, and label-free SPRi-based biosensor for rapid, sensitive and highly selective detection of adenoviruses. The sensing protocol consists of mixing the sample containing adenovirus with a predetermined concentration of adenovirus antibody. The mixture was filtered to remove the free antibodies from the sample. A secondary antibody, which was specific to the adenovirus antibody, was immobilized onto the SPRi chip surface covalently and the filtrate was flowed over the sensor surface. When the free adenovirus antibodies bound to the surface-immobilized secondary antibodies, we observed this binding via changes in reflectivity. In this approach, a higher amount of adenoviruses resulted in fewer free adenovirus antibodies and thus smaller reflectivity changes. A dose-response curve was generated, and the linear detection range was determined to be from 10 PFU/mL to 5000 PFU/mL with an R(2) value greater than 0.9. The results also showed that the developed biosensing system had a high specificity towards adenovirus (less than 20% signal change when tested in a sample matrix containing rotavirus and lentivirus). PMID:26232675

  4. Photoluminescence of double core/shell infrared (CdSeTe)/ZnS quantum dots conjugated to Pseudo rabies virus antibodies

    NASA Astrophysics Data System (ADS)

    Torchynska, T. V.; Casas Espinola, J. L.; Jaramillo Gómez, J. A.; Douda, J.; Gazarian, K.

    2013-06-01

    Double core CdSeTe/ZnS quantum dots (QDs) with emission at 800 nm (1.60 eV) have been studied by photoluminescence (PL) and Raman scattering methods in the non-conjugated state and after the conjugation to the Pseudo rabies virus (PRV) antibodies. The transformation of PL spectra, stimulated by the electric charge of antibodies, has been detected for the bioconjugated QDs. Raman scattering spectra are investigated with the aim to reveal the CdSeTe core compositions. The double core QD energy diagrams were designed that help to analyze the PL spectra and their transformation at the bioconjugation. It is revealed that the interface in double core QDs has the type II quantum well character that permits to explain the near IR optical transition (1.60 eV) in the double core QDs. It is shown that the essential transformation of PL spectra is useful for the study of QD bioconjugation with specific antibodies and can be a powerful technique in early medical diagnostics.

  5. Antibody

    MedlinePlus

    An antibody is a protein produced by the body's immune system when it detects harmful substances, called antigens. Examples ... microorganisms (bacteria, fungi, parasites, and viruses) and chemicals. Antibodies may be produced when the immune system mistakenly ...

  6. Method for double-sided processing of thin film transistors

    DOEpatents

    Yuan, Hao-Chih; Wang, Guogong; Eriksson, Mark A.; Evans, Paul G.; Lagally, Max G.; Ma, Zhenqiang

    2008-04-08

    This invention provides methods for fabricating thin film electronic devices with both front- and backside processing capabilities. Using these methods, high temperature processing steps may be carried out during both frontside and backside processing. The methods are well-suited for fabricating back-gate and double-gate field effect transistors, double-sided bipolar transistors and 3D integrated circuits.

  7. Development and qualification of an antibody rapid deglycosylation method.

    PubMed

    Cook, K Steven; Bullock, Kevin; Sullivan, Timothy

    2012-03-01

    N-linked glycosylation can influence the biological activity and safety of an antibody as well as be a measure of the consistency of the production process. The released N-glycans is an important part of the development of a therapeutic antibody. The traditional method for N-glycan analysis requires complex and laborious sample preparation and lengthy analysis time. Two preparation steps with limited control are removal of the antibody backbone by ice-cold ethanol precipitation and water removal before 2-AB fluorescent dye labeling. Simplification of the sample preparation and better control of key steps that allows for the characterization/quantitation of glycans during all stages of development of a therapeutic antibody is desired. Recently Prozyme introduced a rapid deglycosylation kit and a rapid tagging kit that address some of these issues. The deglycosylation kit immobilizes the antibody on a membrane, thereby eliminating the precipitation step. An instant fluorescent tag kit eliminates the water removal before the 2-AB labeling step. In addition use of a new chromatography column can improve the glycan resolution and shorten the analysis time. The evaluation and qualification of the Rapid Deglycosylation Kit (RDK) and instant 2-AB tagging with the improved chromatography are highlighted. PMID:22257749

  8. Development and Application of a Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Porcine Circovirus 2

    PubMed Central

    Ge, Meng; Luo, Wei; Jiang, Daliang; Li, Runcheng; Zhao, Wenwei; Chen, Guoliang; Yang, Xingdong

    2012-01-01

    A double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) is described for detection of porcine circovirus 2 (PCV2) antibodies using the well-characterized recombinant PCV2 capsid protein. In a comparative test of 394 pig sera against an indirect immunofluorescence (IIF) test and a commercial ELISA kit (also based on the recombinant PCV2 capsid protein), the results showed that the diagnostic sensitivity, specificity, and accuracy of the assay were, respectively, 90.61, 94.02, and 91.62% compared with IIF and 94.38, 95.28, and 94.67% compared with the commercial ELISA kit. Assay of 12 PCV-free pigs over a 5-week period produced only PCV2-negative titers by all 3 methods. These results and the seroprofiles of 4 pig farms obtained by both the commercial ELISA kit and the double-antigen sandwich ELISA indicate that the sandwich ELISA is a reliable method for detection of antibodies to PCV2. Additionally, the method described here permits the use of undiluted test serum samples simultaneously loaded with horseradish peroxidase (HRP)-conjugated antigen into the test well, and the complete test procedure can be performed in less than 90 min. This double-antigen sandwich ELISA should be a useful tool to aid swine industry professionals in deciding the intervention strategies for the control of PCV2-associated diseases. PMID:22815145

  9. Immunogenicity of anti-tumor necrosis factor antibodies-toward improved methods of anti-antibody measurement.

    PubMed

    Aarden, Lucien; Ruuls, Sigrid R; Wolbink, Gertjan

    2008-08-01

    To date, millions of people have been treated with therapeutic monoclonal antibodies (TmAbs) for various indications. It is becoming increasingly clear that TmAbs can be immunogenic, which may reduce efficacy or induce adverse effects. Over the years, the importance of antibody formation has been questioned and sometimes minimized, as few antibody responses to TmAbs (HACA or HAHA) were reported. However, the methods to detect and quantify such antibodies used in the past have been problematic. Only recently, methods have been developed that have adequate sensitivity and are not seriously disturbed by false-positive reactions caused by rheumatoid factors, natural antibodies to Fab or F(ab')2 fragments, or Fc interactions of IgG4. The large number of treated patients, in combination with these new assays, presents a unique opportunity to study the anti-antibody immune response in man, possibly allowing us to manipulate immunogenicity in the future. PMID:18619538

  10. Kinetic of antigent-antibody reactions with scattering method

    NASA Astrophysics Data System (ADS)

    Bilyi, Olexander I.; Kiselyov, Yevgen M.; Novikov, Volodymyr P.

    2001-07-01

    The immune reactions of interaction antigen-antibody represent specific effect of an antigene with an antibody, which outcome are the complex immune aggregates forming precipitate in case of a soluble antigene, or agglutinate in case of a corpuscular antigene. Immunological methods which uses in the quality of carrier protein latex's polymeric microspheresis, gained name and method latex agglutination. Polymeric microspheresis have the array of advantages before biological carries, which consist in the opportunity of the variation of attributes surface and size microspheresis in the broad band of meanings with the preservation of narrow distribution particles behind measurements, the putting of functional groups, necessary for bunch with ligand on stage their synthesis, in ragidity at storage. The quantitative evaluation of parameters of a response of interaction antigen-antibody in immunology is possible by optical methods on a measurement of a modification of intensity of a light stream of a solution in an outcome of a course of a reaction. Concentration of immune complexes determine both on slacking a taking place stream of light, and on a modification of intensity of a stream of light scattering suspended particles in a solution. The process light scattering by colloidal aggregates are formed from suspension microspheresis with adsorbed on their surface protein is described. In report the physics principle of registration immune reaction by light scattering methods is concerned. The results of the effectiveness latex's preparation created on basis of the polymeric carries is described.

  11. Method for limiting heat flux in double-wall tubes

    DOEpatents

    Hwang, Jaw-Yeu

    1982-01-01

    A method of limiting the heat flux in a portion of double-wall tubes including heat treating the tubes so that the walls separate when subjected to high heat flux and supplying an inert gas mixture to the gap at the interface of the double-wall tubes.

  12. Photoluminescence method of testing double heterostructure wafers

    SciTech Connect

    Besomi, P.R.; Wilt, D.P.

    1984-04-10

    Under photoluminescence (PL) excitation, the lateral spreading of photo-excited carriers can suppress the photoluminescence signal from double heterostructure (DH) wafers containing a p-n junction. In any DH with a p-n junction in the active layer, PL is suppressed if the power of the excitation source does not exceed a threshold value. This effect can be advantageously used for a nondestructive optical determination of the top cladding layer sheet conductance as well as p-n junction misplacement, important parameters for injection lasers and LEDs.

  13. A double-sandwich ELISA for identification of monoclonal antibodies suitable for sandwich immunoassays

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The sandwich immunoassay (sIA) is an invaluable technique for concentrating, detecting, and quantifying target antigens. The two critical components required are a capture antibody and a detection antibody, each binding a different epitope on the target antigen. The specific antibodies incorporated...

  14. Occurrence of Newcastle Disease and Infectious Bursal Disease Virus Antibodies in Double-Spurred Francolins in Nigeria

    PubMed Central

    Oluwayelu, Daniel Oladimeji; Adebiyi, Adebowale Idris; Olaniyan, Ibukunoluwa; Ezewele, Phyllis; Aina, Oluwasanmi

    2014-01-01

    The double-spurred francolin Francolinus bicalcaratus has been identified as a good candidate for future domestication due to the universal acceptability of its meat and its adaptability to anthropogenically altered environments. Therefore, in investigating the diseases to which they are susceptible, serum samples from 56 francolins in a major live-bird market (LBM) in Ibadan, southwestern Nigeria, were screened for antibodies against Newcastle disease (ND) and infectious bursal disease (IBD) viruses. Haemagglutination inhibition (HI) test and enzyme-linked immunosorbent assay (ELISA) revealed 25.0% and 35.7% prevalence of ND virus (NDV) antibodies, respectively, while 5.4% and 57.1% prevalence of IBD virus (IBDV) antibodies was detected by agar gel precipitation test (AGPT) and ELISA, respectively. This first report on the occurrence of NDV and IBDV antibodies in apparently healthy, unvaccinated double-spurred francolins from a LBM suggests that they were subclinically infected with either field or vaccine viruses and could thus serve as possible reservoirs of these viruses to domestic poultry. Furthermore, if they are to be domesticated for intensive rearing, a vaccination plan including ND and IBD should be developed and implemented. PMID:26464918

  15. Occurrence of Newcastle Disease and Infectious Bursal Disease Virus Antibodies in Double-Spurred Francolins in Nigeria.

    PubMed

    Oluwayelu, Daniel Oladimeji; Adebiyi, Adebowale Idris; Olaniyan, Ibukunoluwa; Ezewele, Phyllis; Aina, Oluwasanmi

    2014-01-01

    The double-spurred francolin Francolinus bicalcaratus has been identified as a good candidate for future domestication due to the universal acceptability of its meat and its adaptability to anthropogenically altered environments. Therefore, in investigating the diseases to which they are susceptible, serum samples from 56 francolins in a major live-bird market (LBM) in Ibadan, southwestern Nigeria, were screened for antibodies against Newcastle disease (ND) and infectious bursal disease (IBD) viruses. Haemagglutination inhibition (HI) test and enzyme-linked immunosorbent assay (ELISA) revealed 25.0% and 35.7% prevalence of ND virus (NDV) antibodies, respectively, while 5.4% and 57.1% prevalence of IBD virus (IBDV) antibodies was detected by agar gel precipitation test (AGPT) and ELISA, respectively. This first report on the occurrence of NDV and IBDV antibodies in apparently healthy, unvaccinated double-spurred francolins from a LBM suggests that they were subclinically infected with either field or vaccine viruses and could thus serve as possible reservoirs of these viruses to domestic poultry. Furthermore, if they are to be domesticated for intensive rearing, a vaccination plan including ND and IBD should be developed and implemented. PMID:26464918

  16. A Quantitative Method for Comparing the Brightness of Antibody-dye Reagents and Estimating Antibodies Bound per Cell.

    PubMed

    Kantor, Aaron B; Moore, Wayne A; Meehan, Stephen; Parks, David R

    2016-01-01

    We present a quantitative method for comparing the brightness of antibody-dye reagents and estimating antibodies bound per cell. The method is based on complementary binding of test and fill reagents to antibody capture microspheres. Several aliquots of antibody capture beads are stained with varying amounts of the test conjugate. The remaining binding sites on the beads are then filled with a second conjugate containing a different fluorophore. Finally, the fluorescence of the test conjugate compared to the fill conjugate is used to measure the relative brightness of the test conjugate. The fundamental assumption of the test-fill method is that if it takes X molecules of one test antibody to lower the fill signal by Y units, it will take the same X molecules of any other test antibody to give the same effect. We apply a quadratic fit to evaluate the test-fill signal relationship across different amounts of test reagent. If the fit is close to linear, we consider the test reagent to be suitable for quantitative evaluation of antibody binding. To calibrate the antibodies bound per bead, a PE conjugate with 1 PE molecule per antibody is used as a test reagent and the fluorescence scale is calibrated with Quantibrite PE beads. When the fluorescence per antibody molecule has been determined for a particular conjugate, that conjugate can be used for measurement of antibodies bound per cell. This provides comparisons of the brightness of different conjugates when conducted on an instrument whose statistical photoelectron (Spe) scales are known. © 2016 by John Wiley & Sons, Inc. PMID:27367287

  17. A double-antibody sandwich ELISA for the detection of Entamoeba histolytica antigen in stool samples of humans.

    PubMed

    Baumann, D; Gottstein, B

    1987-06-01

    A double-antibody sandwich ELISA was developed to detect detergent-solubilized antigens of Entamoeba histolytica in stool samples of humans. The test system was evaluated for its methodical and diagnostic sensitivity and specificity. In recovery experiments the lower limit of detection was 400 ng E. histolytica (HK9) protein/ml stool, corresponding to approximately 2000 amoebic trophozoites/ml stool. Samples of 97 patients with suspected intestinal amoebiasis were examined. Specific antigens were detected by ELISA (= positive reaction) in 14 (93%) out of 15 stool samples containing trophozoites of E. histolytica. In contrast, 68 (93%) of 73 samples with other protozoa, including Entamoeba coli, E. hartmanni, Endolimax nana, Iodamoeba buetschlii and Giardia lamblia, did not react in the test system (= negative reaction). The test was shown to detect only trophozoites of E. histolytica and not the cyst stage. This fact could facilitate the differentiation between cyst carriers and persons excreting trophozoites. The results of this preliminary study justify a further large scale evaluation of the test system. PMID:2888183

  18. Measuring Double Stars with the Modified Video Drift Method

    NASA Astrophysics Data System (ADS)

    Iverson, Ernest W.; Nugent, Richard L.

    2015-05-01

    The usefulness of a common CCTV video camera for measuring the position angle and separation of a double star is limited by the camera sensitivity and telescope aperture. The video drift method is enhanced by using an integrating camera but frame integrations longer than 0.132 seconds (4 frames) are impractical. This is due to the target stars elongating (streaking) and moving in incremental steps. A simple modification to the Video Drift Method and corresponding VidPro analysis program significantly increases the magnitude at which double stars can be measured. Double stars down to magnitude +16 have been measured with a 14-inch (35.6-cm) telescope using this method compared to magnitude +12 using the original video drift method under comparable seeing conditions.

  19. On 2D bisection method for double eigenvalue problems

    SciTech Connect

    Ji, X.

    1996-06-01

    The two-dimensional bisection method presented in (SIAM J. Matrix Anal. Appl. 13(4), 1085 (1992)) is efficient for solving a class of double eigenvalue problems. This paper further extends the 2D bisection method of full matrix cases and analyses its stability. As in a single parameter case, the 2D bisection method is very stable for the tridiagonal matrix triples satisfying the symmetric-definite condition. Since the double eigenvalue problems arise from two-parameter boundary value problems, an estimate of the discretization error in eigenpairs is also given. Some numerical examples are included. 42 refs., 1 tab.

  20. Proprietary arabinogalactan extract increases antibody response to the pneumonia vaccine: a randomized, double-blind, placebo-controlled, pilot study in healthy volunteers

    PubMed Central

    2010-01-01

    Background Arabinogalactan from Larch tree (Larix spp.) bark has previously demonstrated immunostimulatory activity. The purpose of this study was to test the hypothesis that ingestion of a proprietary arabinogalactan extract, ResistAid™, would selectively enhance the antibody response to the pneumococcal (pneumonia) vaccine in healthy adults. Methods This randomized, double-blind, placebo-controlled, parallel group pilot study included 45 healthy adults who had not previously been vaccinated against Streptococcus pneumoniae. The volunteers began taking the study product or placebo (daily dosage 4.5 g) at the screening visit (V1-Day 0) and continued over the entire 72 day study period. After 30 days the subjects received the 23-valent pneumococcal vaccine (V2). They were monitored the following day (V3-Day 31), as well as 21 days (V4-Day 51) and 42 days (V5-Day 72) after vaccination. Responses by the adaptive immune system (antigen specific) were measured via pneumococcal IgG antibodies (subtypes 4, 6B, 9V, 14, 18C, 19F, and 23F) and salivary IgA levels. Responses by the innate immune system (non-specific) were measured via white blood cell counts, inflammatory cytokines and the complement system. Results Vaccination significantly increased pneumococcal IgG levels as expected. The arabinogalactan group demonstrated a statistically significant greater IgG antibody response than the placebo group in two antibodies subtypes (18C and 23F) at both Day 51 (p = 0.006 and p = 0.002) and at Day 72 (p = 0.008 and p = 0.041). These same subtypes (18C and 23F) also demonstrated change scores from baseline which were significant, in favor of the arabinogalactan group, at Day 51 (p = 0.033 and 0.001) and at Day 72 (p = 0.012 and p = 0.003). Change scores from baseline and mean values were greater in the arabinogalactan group than placebo for most time points in antibody subtypes 4, 6B, 9V, and 19F, but these differences did not reach statistical significance. There was no

  1. An ultrasensitive squamous cell carcinoma antigen biosensing platform utilizing double-antibody single-channel amplification strategy.

    PubMed

    Ren, Xiang; Wu, Dan; Wang, Yuhuan; Zhang, Yunhui; Fan, Dawei; Pang, Xuehui; Li, Yueyun; Du, Bin; Wei, Qin

    2015-10-15

    A novel electrochemical immunosensor was developed for ultrasensitive detection of squamous cell carcinoma antigen (SCCA), which was based on the double-antibody single-channel amplification strategy. For the first time, human immunoglobulin antibody (anti-HIgG) was used as the supporting framework to amplify the loading quantity of SCCA antibody (anti-SCCA). In this strategy, SCCA can be detected without using mesoporous nanometers to amplify the signal. In addition, Pd icosahedrons were first used as the connecter to immobilize the antibodies and strengthen the sensitivity. Only one touch point exists under the limited condition between a sphere and another shape in geometry, thus the Pd icosahedron is an excellent candidate as the role of connecter. Gold nanoparticles (Au NPs) decorated with mercapto-functionalized graphene sheets (Au@GS) were synthesized as the transducing materials. The fabricated immunosensor exhibited an excellent detection limit of 2.8 pg/mL and wide linear range of 0.01-5 ng/mL. This kind of immunosensor would provide a potential application in clinical diagnosis. PMID:25982722

  2. Method for removal of human antibodies to native DNA from serum

    SciTech Connect

    Diamond, B.A.

    1987-09-01

    A method is described for removing human anti-native DNA antibody from a liquid sample comprising coupling monoclonal, antiidiotypic antibodies capable of binding to a shared idiotype on human anti-native DNA antibody to a medium. The idiotype shares between genetically nonidentical individuals, contacting a liquid sample to the medium to permit binding of human anti-native DNA antibody in the sample to the anti-idiotypic antibodies and separating the sample from the medium to remove the human anti-native DNA antibodies therefrom.

  3. Affinity measurement of single chain antibodies: a mathematical method facilitated by statistical software SigmaPlot.

    PubMed

    Safdari, Yaghoub; Farajnia, Safar; Asgharzadeh, Mohammad; Khalili, Masoumeh; Jaliani, Hossein Zarei

    2014-02-01

    Because they are monovalent for antigen, single chain antibodies display a different antibody-antigen interaction pattern from that of full-length antibodies. Using the law of mass action and considering the antibody-antigen binding pattern at OD-100% and OD-50% points, we introduced a formula for estimating single chain antibody affinity. Sigmoid curves of optical density values versus antibody concentrations were drawn and used to determine antibody concentrations at OD-50% points using statistical software SigmaPlot. The OD-50% points were then used to calculate the affinity via the mathematical formula. A software-adapted format of the equation is also presented for further facilitation of the calculation process. The accuracy of this method for affinity calculation was proved by surface plasma resonance. This method offers a precise evaluation of antibody affinity without requiring special material or apparatus, making it possible to be performed in any biological laboratory with minimum facilities. PMID:24555931

  4. [Detection of antigen-antibody interaction of human adenovirus by the method of surface plasmon resonance].

    PubMed

    Nosach, L M; Boltovets', P M; Povnytsia, O Iu; Zhovnovata, V L; Zakharenko, O M; Snopok, B A; Shyrshov, Iu M; Diachenko, N S

    2005-01-01

    A possibility to detect adenoviral protein--hexon, using specific antibodies by surface plasmon resonance (SPR) was demonstrated. The hexon of the human adenovirus 2 (Ad2) binds to antibodies immobilized on the sensor surface treated by KNCS and protein A Staphylococcus aureus. The specificity of antihexon antibodies was demonstrated by indirect method of fluorescent antibodies (MFA) and cellular variant of the immunoassay (cELISA). PMID:16250237

  5. Comparison of double-ended transition state search methods

    NASA Astrophysics Data System (ADS)

    Koslover, Elena F.; Wales, David J.

    2007-10-01

    While a variety of double-ended transition state search methods have been developed, their relative performance in characterizing complex multistep pathways between structurally disparate molecular conformations remains unclear. Three such methods (doubly-nudged elastic band, a string method, and a growing string method) are compared for a series of benchmarks ranging from permutational isomerizations of the seven-atom Lennard-Jones cluster (LJ7) to highly cooperative LJ38 and LJ75 rearrangements, and the folding pathways of two peptides. A database of short paths between LJ13 local minima is used to explore the effects of parameters and suggest reasonable default values. Each double-ended method was employed within the framework of a missing connection network flow algorithm to construct more complicated multistep pathways. We find that in our implementation none of the three methods definitively outperforms the others, and that their relative effectiveness is strongly system and parameter dependent.

  6. Comparison of double-ended transition state search methods.

    PubMed

    Koslover, Elena F; Wales, David J

    2007-10-01

    While a variety of double-ended transition state search methods have been developed, their relative performance in characterizing complex multistep pathways between structurally disparate molecular conformations remains unclear. Three such methods (doubly-nudged elastic band, a string method, and a growing string method) are compared for a series of benchmarks ranging from permutational isomerizations of the seven-atom Lennard-Jones cluster (LJ(7)) to highly cooperative LJ(38) and LJ(75) rearrangements, and the folding pathways of two peptides. A database of short paths between LJ(13) local minima is used to explore the effects of parameters and suggest reasonable default values. Each double-ended method was employed within the framework of a missing connection network flow algorithm to construct more complicated multistep pathways. We find that in our implementation none of the three methods definitively outperforms the others, and that their relative effectiveness is strongly system and parameter dependent. PMID:17919006

  7. Preexisting Antibodies to an F(ab′)2 Antibody Therapeutic and Novel Method for Immunogenicity Assessment

    PubMed Central

    Brady, Ann; Leddy, Cecilia; Coleman, Daniel; Couch, Jessica A.

    2016-01-01

    Anti-therapeutic antibodies (ATAs) may impact drug exposure and activity and induce immune complex mediated toxicity; therefore the accurate measurement of ATA is important for the analysis of drug safety and efficacy. Preexisting ATAs to the hinge region of anti-Delta like ligand 4 (anti-DLL4) F(ab′)2, a potential antitumor therapeutic, were detected in cynomolgus monkey serum, which presented a challenge in developing assays for detecting treatment induced ATA. A total ATA assay was developed using a bridging ELISA that detected both anti-CDR and anti-framework ATA including anti-hinge reactivity. A competition assay that could detect 500 ng/mL of anti-CDR ATA in the presence of preexisting ATA was also developed to determine ATA specific to the anti-DLL4 F(ab′)2 CDR using anti-DLL4 F(ab′)2 and a control F(ab′)2. We used these assay methods in a cynomolgus monkey in vivo study to successfully evaluate total and anti-CDR ATA. The preexisting anti-hinge reactivity was also observed to a lesser extent in human serum, and a similar approach could be applied for specific immunogenicity assessment in clinical trials. PMID:27413757

  8. Method for detecting pathogens attached to specific antibodies

    DOEpatents

    Miles, Robin R.; Venkateswaran, Kodumudi S.; Fuller, Christopher K.

    2005-01-25

    The use of impedance measurements to detect the presence of pathogens attached to antibody-coated beads. In a fluidic device antibodies are immobilized on a surface of a patterned interdigitated electrode. Pathogens in a sample fluid streaming past the electrode attach to the immobilized antibodies, which produces a change in impedance between two adjacent electrodes, which impedance change is measured and used to detect the presence of a pathogen. To amplify the signal, beads coated with antibodies are introduced and the beads would stick to the pathogen causing a greater change in impedance between the two adjacent electrodes.

  9. Double micropipettes configuration method of scanning ion conductance microscopy.

    PubMed

    Zhuang, Jian; Li, Zeqing; Jiao, Yangbohan

    2016-07-01

    In this paper, a new double micropipettes configuration mode of scanning ion conductance microscopy (SICM) is presented to better overcome ionic current drift and further improve the performance of SICM, which is based on a balance bridge circuit. The article verifies the feasibility of this new configuration mode from theoretical and experimental analyses, respectively, and compares the quality of scanning images in the conventional single micropipette configuration mode and the new double micropipettes configuration mode. The experimental results show that the double micropipettes configuration mode of SICM has better effect on restraining ionic current drift and better performance of imaging. Therefore, this article not only proposes a new direction of overcoming the ionic current drift but also develops a new method of SICM stable imaging. PMID:27475561

  10. Double micropipettes configuration method of scanning ion conductance microscopy

    NASA Astrophysics Data System (ADS)

    Zhuang, Jian; Li, Zeqing; Jiao, Yangbohan

    2016-07-01

    In this paper, a new double micropipettes configuration mode of scanning ion conductance microscopy (SICM) is presented to better overcome ionic current drift and further improve the performance of SICM, which is based on a balance bridge circuit. The article verifies the feasibility of this new configuration mode from theoretical and experimental analyses, respectively, and compares the quality of scanning images in the conventional single micropipette configuration mode and the new double micropipettes configuration mode. The experimental results show that the double micropipettes configuration mode of SICM has better effect on restraining ionic current drift and better performance of imaging. Therefore, this article not only proposes a new direction of overcoming the ionic current drift but also develops a new method of SICM stable imaging.

  11. Evaluating method for the double image phenomenon of LED lighting

    NASA Astrophysics Data System (ADS)

    Wu, Wen-Hong; Kuo, Chao-Hui; Hung, Min-Wei; Huang, Kuo-Cheng

    In recent years, the overriding advantages long life, high efficiency, small size and short reaction time have made LED become a viable alternative to conventional light sources. LED lighting sources are usually composed of several individual LED cells which must be mounted on a panel as a lighting module. Being composed of several individual LED cells, the LED sources will cause the double image phenomenon. The double image phenomenon is more obvious when the LED sources are more closer, such as LED table lamp, and limits the applications of LED sources. By using a proper secondary optical lens, the double image phenomenon can be reduced. In this research, an evaluating method based on image processing is developed for the double image phenomenon of a LED sources. By analyzing the gray-scale of the grabbed image which is obtained by putting a rob under a LED source, an index of double image can be established and be a criterion to judge different LED sources. Furthermore, a series of LED lighting simulations are shown in this paper and several type of secondary optical lens are compared and discussed in this paper as well.

  12. Double wall vacuum tubing and method of manufacture

    DOEpatents

    Stahl, Charles R.; Gibson, Michael A.; Knudsen, Christian W.

    1989-01-01

    An evacuated double wall tubing is shown together with a method for the manufacture of such tubing which includes providing a first pipe of predetermined larger diameter and a second pipe having an O.D. substantially smaller than the I.D. of the first pipe. An evacuation opening is then in the first pipe. The second pipe is inserted inside the first pipe with an annular space therebetween. The pipes are welded together at one end. A stretching tool is secured to the other end of the second pipe after welding. The second pipe is then prestressed mechanically with the stretching tool an amount sufficient to prevent substantial buckling of the second pipe under normal operating conditions of the double wall pipe. The other ends of the first pipe and the prestressed second pipe are welded together, preferably by explosion welding, without the introduction of mechanical spacers between the pipes. The annulus between the pipes is evacuated through the evacuation opening, and the evacuation opening is finally sealed. The first pipe is preferably of steel and the second pipe is preferably of titanium. The pipes may be of a size and wall thickness sufficient for the double wall pipe to be structurally load bearing or may be of a size and wall thickness insufficient for the double wall pipe to be structurally load bearing, and the double wall pipe positioned with a sliding fit inside a third pipe of a load-bearing size.

  13. Development of a double-monoclonal antibody sandwich ELISA: Tool for chicken interferon-γ detection ex vivo.

    PubMed

    Dai, Hua; Xu, Zheng-Zhong; Wang, Meiling; Chen, Jun-Hua; Chen, Xiang; Pan, Zhi-Ming; Jiao, Xin-An

    2016-04-01

    The aim of the present work was to develop reagents to set up a chicken interferon-γ (ChIFN-γ) assay. Four monoclonal antibodies (mAbs) specific for ChIFN-γ were generated to establish sandwich ELISA based on 2 different mAbs. To improve the detection sensitivity of ChIFN-γ, a double-monoclonal antibody sandwich ELISA was developed using mAb 3E5 as capture antibody and biotinylated mAb 3E3 as a detection reagent. The results revealed that this ELISA has high sensitivity, allowing for the detection of 125 to 500 pg/mL of recombinant ChIFN-γ, and also has an excellent capacity for detecting native ChIFN-γ. This ELISA was then used to detect ChIFN-γ level in chickens immunized with a Newcastle disease virus (NDV) vaccine, the immunized chicken splenocytes were stimulated by NDV F protein as recall antigen. From our results, it appears that the sensitivity range of this sandwich ELISA test is adequate to measure the ex vivo release of ChIFN-γ. PMID:27127340

  14. Concerning the Video Drift Method to Measure Double Stars

    NASA Astrophysics Data System (ADS)

    Nugent, Richard L.; Iverson, Ernest W.

    2015-05-01

    Classical methods to measure position angles and separations of double stars rely on just a few measurements either from visual observations or photographic means. Visual and photographic CCD observations are subject to errors from the following sources: misalignments from eyepiece/camera/barlow lens/micrometer/focal reducers, systematic errors from uncorrected optical distortions, aberrations from the telescope system, camera tilt, magnitude and color effects. Conventional video methods rely on calibration doubles and graphically calculating the east-west direction plus careful choice of select video frames stacked for measurement. Atmospheric motion is one of the larger sources of error in any exposure/measurement method which is on the order of 0.5-1.5. Ideally, if a data set from a short video can be used to derive position angle and separation, with each data set self-calibrating independent of any calibration doubles or star catalogues, this would provide measurements of high systematic accuracy. These aims are achieved by the video drift method first proposed by the authors in 2011. This self calibrating video method automatically analyzes 1,000's of measurements from a short video clip.

  15. Conjugation of R-Phycoerythrin to a Polyclonal Antibody and F (ab')2 Fragment of a Polyclonal Antibody by Two Different Methods.

    PubMed

    Mahmoudian, Jafar; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah; Mahmoudi, Ahmad Reza; Akhondi, Mohammad Mehdi; Zarnani, Amir Hassan; Goli, Leila Balaei; Babaei, Mahdokht; Ghods, Roya

    2010-04-01

    R-Phycoerythrin (R-PE), a fluorescent protein from phycobiliprotein family, is isolated from red algae. Conjugation of antibodies to R-PE facilitates multiple fluorescent staining methods. In the present study polyclonal antibodies and polyclonal F(ab')2 fragment antibodies were conjugated to R-PE by two different methods. The efficiency of the methods was evaluated using Immunocytochemistry (ICC) and Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE). In the first conjugation method, PE was attached to SMCC linker followed by conjugation of antibody to PE-SMCC. In the second method, SH groups were added onto R-PE molecule, while the antibody was attached to SPDP linker. Then, the antibody-SPDP molecule was conjugated to R-PE. Our results showed that the two conjugation methods did not have any abrogative effects on the antibody binding activity. PMID:23407609

  16. Differentiation between Human Coronaviruses NL63 and 229E Using a Novel Double-Antibody Sandwich Enzyme-Linked Immunosorbent Assay Based on Specific Monoclonal Antibodies

    PubMed Central

    Sastre, Patricia; Dijkman, Ronald; Camuñas, Ana; Ruiz, Tamara; Jebbink, Maarten F.; van der Hoek, Lia; Vela, Carmen; Rueda, Paloma

    2011-01-01

    Human coronaviruses (HCoVs) are responsible for respiratory tract infections ranging from common colds to severe acute respiratory syndrome. HCoV-NL63 and HCoV-229E are two of the four HCoVs that circulate worldwide and are close phylogenetic relatives. HCoV infections can lead to hospitalization of children, elderly individuals, and immunocompromised patients. Globally, approximately 5% of all upper and lower respiratory tract infections in hospitalized children are caused by HCoV-229E and HCoV-NL63. The latter virus has recently been associated with the childhood disease croup. Thus, differentiation between the two viruses is relevant for epidemiology studies. The aim of this study was to develop a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) as a potential tool for identification and differentiation between HCoV-NL63 and HCoV-229E. The nucleocapsid (N) proteins of HCoV-NL63 and HCoV-229E were expressed in an Escherichia coli system and used to immunize mice in order to obtain monoclonal antibodies (MAbs) specific for each virus. Three specific MAbs to HCoV-NL63, one MAb specific to HCoV-229E, and four MAbs that recognized both viruses were obtained. After their characterization, three MAbs were selected in order to develop a differential DAS-ELISA. The described assay could detect up to 3 ng/ml of N protein and 50 50% tissue culture infective doses/ml of virus stock. No cross-reactivity with other human coronaviruses or closely related animal coronaviruses was found. The newly developed DAS-ELISA was species specific, and therefore, it could be considered a potential tool for detection and differentiation of HCoV-NL63 and HCoV-229E infections. PMID:21084464

  17. Differentiation between human coronaviruses NL63 and 229E using a novel double-antibody sandwich enzyme-linked immunosorbent assay based on specific monoclonal antibodies.

    PubMed

    Sastre, Patricia; Dijkman, Ronald; Camuñas, Ana; Ruiz, Tamara; Jebbink, Maarten F; van der Hoek, Lia; Vela, Carmen; Rueda, Paloma

    2011-01-01

    Human coronaviruses (HCoVs) are responsible for respiratory tract infections ranging from common colds to severe acute respiratory syndrome. HCoV-NL63 and HCoV-229E are two of the four HCoVs that circulate worldwide and are close phylogenetic relatives. HCoV infections can lead to hospitalization of children, elderly individuals, and immunocompromised patients. Globally, approximately 5% of all upper and lower respiratory tract infections in hospitalized children are caused by HCoV-229E and HCoV-NL63. The latter virus has recently been associated with the childhood disease croup. Thus, differentiation between the two viruses is relevant for epidemiology studies. The aim of this study was to develop a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) as a potential tool for identification and differentiation between HCoV-NL63 and HCoV-229E. The nucleocapsid (N) proteins of HCoV-NL63 and HCoV-229E were expressed in an Escherichia coli system and used to immunize mice in order to obtain monoclonal antibodies (MAbs) specific for each virus. Three specific MAbs to HCoV-NL63, one MAb specific to HCoV-229E, and four MAbs that recognized both viruses were obtained. After their characterization, three MAbs were selected in order to develop a differential DAS-ELISA. The described assay could detect up to 3 ng/ml of N protein and 50 50% tissue culture infective doses/ml of virus stock. No cross-reactivity with other human coronaviruses or closely related animal coronaviruses was found. The newly developed DAS-ELISA was species specific, and therefore, it could be considered a potential tool for detection and differentiation of HCoV-NL63 and HCoV-229E infections. PMID:21084464

  18. Double sided circuit board and a method for its manufacture

    DOEpatents

    Lindenmeyer, C.W.

    1988-04-14

    Conductance between the sides of a large double sided printed circuit board is provided using a method which eliminates the need for chemical immersion or photographic exposure of the entire large board. A plurality of through-holes are drilled or punched in a substratum according to the desired pattern, conductive laminae are made to adhere to both sides of the substratum covering the holes and the laminae are pressed together and permanently joined within the holes, providing conductive paths. 4 figs.

  19. Double sided circuit board and a method for its manufacture

    DOEpatents

    Lindenmeyer, Carl W.

    1989-01-01

    Conductance between the sides of a large double sided printed circuit board is provided using a method which eliminates the need for chemical immersion or photographic exposure of the entire large board. A plurality of through-holes are drilled or punched in a substratum according to the desired pattern, conductive laminae are made to adhere to both sides of the substratum covering the holes and the laminae are pressed together and permanently joined within the holes, providing conductive paths.

  20. Double sided circuit board and a method for its manufacture

    DOEpatents

    Lindenmeyer, Carl W.

    1989-07-04

    Conductance between the sides of a large double sided printed circuit board is provided using a method which eliminates the need for chemical immersion or photographic exposure of the entire large board. A plurality of through-holes are drilled or punched in a substratum according to the desired pattern, conductive laminae are made to adhere to both sides of the substratum covering the holes and the laminae are pressed together and permanently joined within the holes, providing conductive paths.

  1. Heart antibodies in cardiomyopathies.

    PubMed Central

    Trueman, T; Thompson, R A; Cummins, P; Littler, W A

    1981-01-01

    The reported frequency of circulating heart reactive antibodies in cardiomyopathies has varied and their significance is unknown. In this study such antibodies were sought in patients with primary congestive and hypertrophic cardiomyopathies and other heart diseases. Standard "single sandwich" and the more sensitive "double sandwich" indirect immunofluorescence techniques failed to disclose a significant difference between any cardiomyopathic group and controls in repeated experiments. With both techniques results were subject to considerable method-specific artefacts and observer variation. No published work associating heart antibodies detected by immunofluorescence methods with cariomyopathies adequately takes these into account. PMID:7028058

  2. Characterization of immobilization methods of antiviral antibodies in serum for electrochemical biosensors

    NASA Astrophysics Data System (ADS)

    Huy, Tran Quang; Hanh, Nguyen Thi Hong; Van Chung, Pham; Anh, Dang Duc; Nga, Phan Thi; Tuan, Mai Anh

    2011-06-01

    In this paper, we describes different methods to immobilize Japanese encephalitis virus (JEV) antibodies in human serum onto the interdigitated surface of a microelectrode sensor for optimizing electrochemical detection: (1) direct covalent binding to the silanized surface, (2) binding to the silanized surface via a cross-linker of glutaraldehyde (GA), (3) binding to glutaraldehyde/silanized surface via goat anti-human IgG polyclonal antibody and (4) binding to glutaraldehyde/silanized surface via protein A (PrA). Field emission scanning electron microscopy, Fourier transform infrared spectrometry, and fluorescence microscopy are used to verify the characteristics of antibodies on the interdigitated surface after the serum antibodies immobilization. The analyzed results indicate that the use of protein A is an effective choice for immobilization and orientation of antibodies in serum for electrochemical biosensors. This study provides an advantageous immobilization method of serum containing antiviral antibodies to develop electrochemical biosensors for preliminary screening of viruses in clinical samples from outbreaks.

  3. A method for generating double-ring-shaped vector beams

    NASA Astrophysics Data System (ADS)

    Huan, Chen; Xiao-Hui, Ling; Zhi-Hong, Chen; Qian-Guang, Li; Hao, Lv; Hua-Qing, Yu; Xu-Nong, Yi

    2016-07-01

    We propose a method for generating double-ring-shaped vector beams. A step phase introduced by a spatial light modulator (SLM) first makes the incident laser beam have a nodal cycle. This phase is dynamic in nature because it depends on the optical length. Then a Pancharatnam–Berry phase (PBP) optical element is used to manipulate the local polarization of the optical field by modulating the geometric phase. The experimental results show that this scheme can effectively create double-ring-shaped vector beams. It provides much greater flexibility to manipulate the phase and polarization by simultaneously modulating the dynamic and the geometric phases. Project supported by the National Natural Science Foundation of China (Grant No. 11547017), the Hubei Engineering University Research Foundation, China (Grant No. z2014001), and the Natural Science Foundation of Hubei Province, China (Grant No. 2014CFB578).

  4. Double-Antigen Enzyme-Linked Immunosorbent Assay for Detection of Hepatitis E Virus-Specific Antibodies in Human or Swine Sera ▿

    PubMed Central

    Hu, Wei Ping; Lu, Yang; Precioso, Nestor Amadeo; Chen, Hsiao Ying; Howard, Teresa; Anderson, David; Guan, Ming

    2008-01-01

    A new double-antigen sandwich-based enzyme-linked immunosorbent assay (ELISA) for the detection of total antibodies (immunoglobulin G [IgG] and IgM) specific for hepatitis E virus (HEV) was developed by utilizing well-characterized recombinant protein ET2.1 and its peroxidase-labeled counterpart. Our study showed that the ELISA detected all the positive patient samples (n = 265) regardless of whether they contained IgM or IgG antibodies, or both, while it maintained an excellent specificity of 98.8% with samples from various patient or healthy control groups (total number of samples, 424). The test had a detection limit for anti-HEV IgG antibodies that was equivalent to 62 mIU/ml of the international reference. Compared with the serological status of the specimens determined on the basis of tests performed at the individual collection sites, the testing outcome generated by the new ELISA had a good agreement of 99.3%, with a kappa value of 0.985. The positive predictive value and the negative predictive value for the new test reached 98.1% and 100%, respectively. This ELISA had a positive delta value of 4.836 and a negative delta value of 3.314 (where delta is a measure of the number of standard deviations by which the cutoff is separated from the mean of the sample groups) (N. Crofts, W. Maskill, and I. D. Gust, J. Virol. Methods 22:51-59, 1988), indicating that it had an excellent ability to differentiate the infected and noninfected cohorts. Furthermore, the new design enables the detection of antibodies not only in human samples but also in pig samples. Our preliminary data showed that the ELISA could detect seroconversion in samples from pigs at as early as 14 days postinoculation. The potential utility of detecting specific antibodies in pigs will be an added advantage for managing the disease, with suggested zoonotic implications. PMID:18495846

  5. A fluorescence-based method to directly quantify antibodies immobilized on gold nanoparticles.

    PubMed

    Filbrun, Seth L; Driskell, Jeremy D

    2016-06-21

    The ability to evaluate antibody immobilization onto gold nanoparticles is critical for assessing coupling chemistry and optimizing the sensitivity of nanoparticle-enabled biosensors. Herein, we developed a fluorescence-based method for directly quantifying antibodies bound onto gold nanoparticles. Antibody-modified gold nanoparticles were treated with KI/I2 etchant to dissolve the gold nanoparticles. A desalting spin column was used to recover the antibody released from the nanoparticles, and NanoOrange, a fluorescent dye, was used to quantify the antibody. We determined 309 ± 93 antibodies adsorb onto a 60 nm gold nanoparticles (2.6 × 10(10) NP mL(-1)), which is consistent with a fully adsorbed monolayer based on the footprint of an IgG molecule. Moreover, the increase in hydrodynamic diameter of the conjugated nanoparticle (76 nm) compared to that of the unconjugated nanoparticle (62 nm) confirmed that multilayers did not form. A more conventional method of indirectly quantifying the adsorbed antibody by analysis of the supernatant overestimated the antibody surface coverage (660 ± 87 antibodies per nanoparticle); thus we propose the method described herein as a more accurate alternative to the conventional approach. PMID:27113720

  6. Machine learning methods enable predictive modeling of antibody feature:function relationships in RV144 vaccinees.

    PubMed

    Choi, Ickwon; Chung, Amy W; Suscovich, Todd J; Rerks-Ngarm, Supachai; Pitisuttithum, Punnee; Nitayaphan, Sorachai; Kaewkungwal, Jaranit; O'Connell, Robert J; Francis, Donald; Robb, Merlin L; Michael, Nelson L; Kim, Jerome H; Alter, Galit; Ackerman, Margaret E; Bailey-Kellogg, Chris

    2015-04-01

    The adaptive immune response to vaccination or infection can lead to the production of specific antibodies to neutralize the pathogen or recruit innate immune effector cells for help. The non-neutralizing role of antibodies in stimulating effector cell responses may have been a key mechanism of the protection observed in the RV144 HIV vaccine trial. In an extensive investigation of a rich set of data collected from RV144 vaccine recipients, we here employ machine learning methods to identify and model associations between antibody features (IgG subclass and antigen specificity) and effector function activities (antibody dependent cellular phagocytosis, cellular cytotoxicity, and cytokine release). We demonstrate via cross-validation that classification and regression approaches can effectively use the antibody features to robustly predict qualitative and quantitative functional outcomes. This integration of antibody feature and function data within a machine learning framework provides a new, objective approach to discovering and assessing multivariate immune correlates. PMID:25874406

  7. A radioimmunoassay method for detection of DNA based on chemical immobilization of anti-DNA antibody.

    PubMed

    Yoo, S K; Yoon, M; Park, U J; Han, H S; Kim, J H; Hwang, H J

    1999-09-30

    High selectivity provided by biomolecules such as antibodies and enzymes has been exploited during the last two decades for development of biosensors. Of particular importance are efficient immobilization methods for biomolecules in order to preserve their biological activities. In this study, we have evaluated immobilization strategies for an anti-DNA antibody on a self-assembled monolayer of omega-functionalized thiols. The antibody was immobilized via peptide bond formation between the primary amines in the antibody and the carboxyl groups on the self-assembled monolayer. The peptide bond coupling was achieved by activating COOH groups on the surface through N-Hydroxysuccimide (NHS)-ester formation, followed by acylation of NH2 group in the antibody. DNA binding activity of the immobilized antibody was examined by counting beta emission from 35S-labeled DNA. PMID:10551259

  8. Monoclonal antibodies to cyclodiene insecticides and method for detecting the same

    DOEpatents

    Stanker, L.H.; Vanderlaan, M.; Watkins, B.E.

    1994-08-02

    Methods are described for making specific monoclonal antibodies useful for detection of cyclodienes in foods and environmental samples. Monoclonal antibodies specifically reactive with cyclodienes can detect accumulated pesticides in food, tissue or environmental samples. Extraction and preparation of organic samples for immunoassay in a polar-nonpolar reaction medium permits detection of halogenated organic ring structures at concentrations in samples. 13 figs.

  9. Monoclonal antibodies to cyclodiene insecticides and method for detecting the same

    DOEpatents

    Stanker, Larry H.; Vanderlaan, Martin; Watkins, Bruce E.

    1994-01-01

    Methods are described for making specific monoclonal antibodies useful for detection of cyclodienes in foods and environmental samples. Monoclonal antibodies specifically reactive with cyclodienes can detect accumulated pesticides in food, tissue or environmental samples. Extraction and preparation of organic samples for immunoassay in a polar-nonpolar reaction medium permits detection of halogenated organic ring structures at concentrations in samples.

  10. Method of rapid production of hybridomas expressing monoclonal antibodies on the cell surface

    DOEpatents

    Meagher, Richard B.; Laterza, Vince

    2006-12-12

    The present invention relates to genetically altered hybridomas, myelomas and B cells. The invention also relates to utilizing genetically altered hybridomas, myelomas and B cells in methods of making monoclonal antibodies. The present invention also provides populations of hybridomas and B cells that can be utilized to make a monoclonal antibody of interest.

  11. New Methods for Improved Double Circular-Arc Helical Gears

    NASA Technical Reports Server (NTRS)

    Litvin, Faydor L.; Lu, Jian

    1997-01-01

    The authors have extended the application of double circular-arc helical gears for internal gear drives. The geometry of the pinion and gear tooth surfaces has been determined. The influence of errors of alignment on the transmission errors and the shift of the bearing contact have been investigated. Application of a predesigned parabolic function for the reduction of transmission errors was proposed. Methods of grinding of the pinion-gear tooth surfaces by a disk-shaped tool and a grinding worm were proposed.

  12. A fluorescent imaging method for analyzing the biodistribution of therapeutic monoclonal antibodies that can distinguish intact antibodies from their breakdown products

    PubMed Central

    Suzuki, Takuo; Miyazaki, Chihiro; Ishii-Watabe, Akiko; Tada, Minoru; Sakai-Kato, Kumiko; Kawanishi, Toru; Kawasaki, Nana

    2015-01-01

    Many monoclonal antibodies have been developed for therapy over the last 2 decades. In the development of therapeutic antibodies, the preclinical assessment of an antibody's biodistribution is important for the prediction of the antibody's efficacy and safety. For imaging analyses of such biodistributions, radioisotope (RI) labeling and fluorescence labeling methods are typically used, but the resulting data are limited because these methods cannot distinguish breakdown products from intact antibodies. To resolve this problem, we developed a novel method using fluorescent resonance energy transfer (FRET)-type labeling and a spectral unmixing tool. With FRET-type labeling (labeling with 2 species of fluorophore), different fluorescence properties of labeled intact antibodies and their breakdown products (the hydrolyzed/digested type of breakdown products) are made visible. With the spectral unmixing tool, the fluorescence of a solution containing the intact antibody and its breakdown products could be unmixed in proportion to their contents. Moreover, when labeled antibodies that targeted either human epidermal growth factor receptor-2 or epidermal growth factor receptor were injected into nude mice implanted subcutaneously with tumor cells, the accumulation of the injected labeled antibodies and their breakdown products in the tumor could be separately analyzed by both whole-mouse imaging and a tumor homogenate analysis. These results suggest that our method using FRET-type labeling and a spectral unmixing tool could be useful in distinguishing breakdown products from intact antibodies. PMID:25891896

  13. A Novel Antibody Humanization Method Based on Epitopes Scanning and Molecular Dynamics Simulation

    PubMed Central

    Zhao, Bin-Bin; Gong, Lu-Lu; Jin, Wen-Jing; Liu, Jing-Jun; Wang, Jing-Fei; Wang, Tian-Tian; Yuan, Xiao-Hui; He, You-Wen

    2013-01-01

    1-17-2 is a rat anti-human DEC-205 monoclonal antibody that induces internalization and delivers antigen to dendritic cells (DCs). The potentially clinical application of this antibody is limited by its murine origin. Traditional humanization method such as complementarity determining regions (CDRs) graft often leads to a decreased or even lost affinity. Here we have developed a novel antibody humanization method based on computer modeling and bioinformatics analysis. First, we used homology modeling technology to build the precise model of Fab. A novel epitope scanning algorithm was designed to identify antigenic residues in the framework regions (FRs) that need to be mutated to human counterpart in the humanization process. Then virtual mutation and molecular dynamics (MD) simulation were used to assess the conformational impact imposed by all the mutations. By comparing the root-mean-square deviations (RMSDs) of CDRs, we found five key residues whose mutations would destroy the original conformation of CDRs. These residues need to be back-mutated to rescue the antibody binding affinity. Finally we constructed the antibodies in vitro and compared their binding affinity by flow cytometry and surface plasmon resonance (SPR) assay. The binding affinity of the refined humanized antibody was similar to that of the original rat antibody. Our results have established a novel method based on epitopes scanning and MD simulation for antibody humanization. PMID:24278299

  14. Biotin radioligand assay with an /sup 125/I-labeled biotin derivative, avidin, and avidin double-antibody reagents

    SciTech Connect

    Livaniou, E.; Evangelatos, G.P.; Ithakissios, D.S.

    1987-11-01

    We describe a new radioligand assay for determining biotin in biological fluids by using a mixture of N-(beta-(4-OH-3-125I-phenyl)ethyl)- and N-(beta-(4-OH-3,5-di-125I-phenyl)ethyl)biotinamides as radiotracer, avidin as a binding protein, and an avidin double-antibody as a separation reagent. The radiotracer is synthesized by coupling (at pH 8.5, 20-22 degrees C, 90 min) N-hydroxysuccinimidobiotin to radioiodinated tyramine. The assay curve is linear and the assay itself is sensitive (less than 10 ng/L), reproducible (intra- and interassay CVs 4.1% and 7.0%, respectively), and allows the simultaneous handling of more than 100 samples in less than 4 h. Serum samples from apparently normal subjects contained 100-840 ng of biotin per liter (mean 340 ng/L). Pregnant women had low concentrations of biotin (100-300 ng/L) in their serum. Patients undergoing chronic hemodialysis treatment showed high concentrations (0.5-3.0 micrograms/L), which may be ascribable to the inability of avidin, which was used as the assay binding protein, to distinguish biotin from biotinyl derivatives with an intact ureido ring.

  15. Antinuclear antibodies and their detection methods in diagnosis of connective tissue diseases: a journey revisited

    PubMed Central

    Kumar, Yashwant; Bhatia, Alka; Minz, Ranjana Walker

    2009-01-01

    It has been more than 50 years since antinuclear antibodies were first discovered and found to be associated with connective tissue diseases. Since then different methods have been described and used for their detection or confirmation. For many decades immunofluorescent antinuclear antibody test has been the "gold standard" in the diagnosis of these disorders. However to increase the sensitivity and specificity of antinuclear antibody detection further approaches were explored. Today a battery of newer techniques are available some of which are now considered better and are competing with the older methods. This article provides an overview on advancement in antinuclear antibody detection methods, their future prospects, advantages, disadvantages and guidelines for use of these tests. PMID:19121207

  16. Monoclonal antibodies for the separate detection of halodeoxyuridines and method for their use

    DOEpatents

    Vanderlaan, Martin; Watkins, Bruce E.; Stanker, Larry H.

    1991-01-01

    Monoclonal antibodies are described which have specific affinities for halogenated nucleoside analogs and are preferentially selective for one particular halogen. Such antibodies, when incorporated into immunochemical reagents, may be used to identify and independently quantify the cell division character of more than one population or subpopulation in flow cytometric measurements. Independent assessment of division activity in cell sub-populations facilitates selection of appropriate time and dose for administration of anti-proliferative agents. The hybridomas which secrete halogen selective antibodies and the method of making them are described.

  17. Monoclonal antibodies for the separate detection of halodeoxyuridines and method for their use

    DOEpatents

    Vanderlaan, M.; Watkins, B.E.; Stanker, L.H.

    1991-10-01

    Monoclonal antibodies are described which have specific affinities for halogenated nucleoside analogs and are preferentially selective for one particular halogen. Such antibodies, when incorporated into immunochemical reagents, may be used to identify and independently quantify the cell division character of more than one population or subpopulation in flow cytometric measurements. Independent assessment of division activity in cell sub-populations facilitates selection of appropriate time and dose for administration of anti-proliferative agents. The hybridomas which secrete halogen selective antibodies and the method of making them are described. 14 figures.

  18. A simple method for the production of anti-C3d monoclonal antibody.

    PubMed

    Cruz, Carlos; León, Graciela

    2007-12-01

    Production of monoclonal antibodies to C3d usually involves the purification of protein. Our method does not require C3 purification; it relies on attachment of C3b to mouse erythrocytes by activation of alternative pathways and further conversion in C3d. We prepared human complement-coated mouse red cells and sensitized mice of the same strain with our own schedule of immunization and applied the classical methods to obtain a mouse monoclonal antibody. We obtained a clone called BMS-11 which produces a monoclonal antibody of IgM class, to C3d with a title of 1:500000. The monoclonal antibody obtained has shown that it is suitable for use as an antiglobulin reagent. PMID:18158789

  19. Serotype- and serogroup-specific detection of African horsesickness virus using phage displayed chicken scFvs for indirect double antibody sandwich ELISAs.

    PubMed

    van Wyngaardt, Wouter; Mashau, Cordelia; Wright, Isabel; Fehrsen, Jeanni

    2013-01-01

    There is an ongoing need for standardized, easily renewable immunoreagents for detecting African horsesickness virus (AHSV). Two phage displayed single-chain variable fragment (scFv) antibodies, selected from a semi-synthetic chicken antibody library, were used to develop double antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) to detect AHSV. In the DAS-ELISAs, the scFv previously selected with directly immobilized AHSV-3 functioned as a serotype-specific reagent that recognized only AHSV-3. In contrast, the one selected with AHSV-8 captured by IgG against AHSV-3 recognized all nine AHSV serotypes but not the Bryanston strain of equine encephalosis virus. Serving as evidence for its serogroup-specificity. These two scFvs can help to rapidly confirm the presence of AHSV while additional serotype-specific scFvs may simplify AHSV serotyping. PMID:23388433

  20. Upscaling of Hydraulic Conductivity using the Double Constraint Method

    NASA Astrophysics Data System (ADS)

    El-Rawy, Mustafa; Zijl, Wouter; Batelaan, Okke

    2013-04-01

    The mathematics and modeling of flow through porous media is playing an increasingly important role for the groundwater supply, subsurface contaminant remediation and petroleum reservoir engineering. In hydrogeology hydraulic conductivity data are often collected at a scale that is smaller than the grid block dimensions of a groundwater model (e.g. MODFLOW). For instance, hydraulic conductivities determined from the field using slug and packer tests are measured in the order of centimeters to meters, whereas numerical groundwater models require conductivities representative of tens to hundreds of meters of grid cell length. Therefore, there is a need for upscaling to decrease the number of grid blocks in a groundwater flow model. Moreover, models with relatively few grid blocks are simpler to apply, especially when the model has to run many times, as is the case when it is used to assimilate time-dependent data. Since the 1960s different methods have been used to transform a detailed description of the spatial variability of hydraulic conductivity to a coarser description. In this work we will investigate a relatively simple, but instructive approach: the Double Constraint Method (DCM) to identify the coarse-scale conductivities to decrease the number of grid blocks. Its main advantages are robustness and easy implementation, enabling to base computations on any standard flow code with some post processing added. The inversion step of the double constraint method is based on a first forward run with all known fluxes on the boundary and in the wells, followed by a second forward run based on the heads measured on the phreatic surface (i.e. measured in shallow observation wells) and in deeper observation wells. Upscaling, in turn is inverse modeling (DCM) to determine conductivities in coarse-scale grid blocks from conductivities in fine-scale grid blocks. In such a way that the head and flux boundary conditions applied to the fine-scale model are also honored at the

  1. Production of monoclonal antibodies specific for Haemophilus ducreyi: a screening method to discriminate specific and cross-reacting antibodies.

    PubMed

    Odumeru, J A; Alfa, M J; Martin, C F; Ronald, A R; Jay, F T

    1989-06-01

    Haemophilus ducreyi is the etiological agent of chancroid. The organism shares extensive immunological cross-reactivity with other Haemophilus species. This presents substantial difficulties for the production of specific monoclonal antibodies (MAbs). A competition ELISA was devised for hybridoma screening which allowed the detection of H. ducreyi-specific antibody-producing hybridoma cultures during the initial screening process. With this screening method, seven MAbs specific for H. ducreyi were obtained in a single cell fusion exercise. The specificities of the 7 MAbs were demonstrated by direct ELISA and dot immunobinding assays against several strains each of H. influenzae, H. parainfluenzae and Neisseria gonorrhoeae. Five of the MAbs reacted against all ten strains of H. ducreyi. These MAbs may permit the development of rapid and efficient immunodiagnostics for chancroid. The principle of the competition ELISA for hybridoma screening should be widely applicable to the development of specific MAbs to other organisms in which immunological cross-reactivity is an impediment to hybridoma screening by conventional methods. PMID:2787274

  2. Antineutrophil cytoplasmic antibodies (ANCA) testing: detection methods and clinical application.

    PubMed

    Sinico, Renato Alberto; Radice, Antonella

    2014-01-01

    Antineutrophil cytoplasmic antibodies (ANCA) are considered the diagnostic biomarker of some necrotising vasculitis such as granulomatosis with polyangiitis (GPA), microscopic polyangiitis (MPA) and, to a lesser extent, eosinophilic granulomatosis with polyangiitis (EGPA). According to the current recommendations, combining indirect immunofluorescence and proteinase 3 (PR3) and myeloperoxidase (MPO) antigen specific immunometric assays, in the proper clinical setting, assures the best diagnostic specificity. When such conditions are satisfied, ANCA are detected in up to 90% of patients with active generalised GPA and MPA and in about 40% of patients with EGPA. Cytoplasmic ANCA (C-ANCA) with specificity for PR3 are usually found in patients with GPA whereas perinuclear ANCA (P-ANCA) in patients with MPA and EGPA. However, ANCA antigen specificity is more closely associated with disease phenotype and prognosis than clinical diagnosis. The clinical value of serial ANCA testing in monitoring disease activity is still debated. Recently, new promising developments in methodology and techniques (computer-based image analysis of immunofluorescence patterns, novel generation of PR3-/MPO-ANCA immunometric assays and multiplex technology) have been proposed but studies comparing the performances of the different assays are scarce. PMID:24854381

  3. Diagnostic methods for African horsesickness virus using monoclonal antibodies to structural and non-structural proteins.

    PubMed

    Ranz, A I; Miguet, J G; Anaya, C; Venteo, A; Cortés, E; Vela, C; Sanz, A

    1992-11-01

    A panel of 32 hybridoma cell lines secreting monoclonal antibodies (MAbs) reactive with African horsesickness virus serotype 4 (AHSV-4) has been developed. Four of the MAbs recognized the major core antigen VP7, twenty recognized the outer capsid protein VP2 and eight reacted with the non-structural protein NS1. With the VP7-specific MAbs a rapid and sensitive double antibody sandwich immunoassay has been developed to detect viral antigen in infected Vero cells and in spleen tissue from AHSV-infected horses. The sensitivity of the assay is 10 ng viral antigen per 100 microliters. The NS1-specific MAbs allowed visualization by immunofluorescence of tubule-like structures in the cytoplasm of infected Vero cells. This can be very useful as a confirmatory diagnostic procedure. The antigenic map of the outer capsid VP2 protein with MAbs is also reported. PMID:1481354

  4. [The development of methods for obtaining monoclonal antibody-producing cells].

    PubMed

    Skowicki, Michał; Lipiński, Tomasz

    2016-01-01

    Monoclonal antibodies (mAbs) are biomolecules of great scientific and practical significance. In contrast to polyclonal antibodies from immune sera, they are homogeneous and monospecific, since they are produced by hybridoma cells representing a clone arising from a single cell. The successful technology was described for the first time in 1975; the inventors were later awarded the Nobel Prize. Currently, mAbs are broadly used as a research tool, in diagnostics and medicine in particular for the treatment of cancer or in transplantology. About 47 therapeutics based on monoclonal antibodies are now available in the US and Europe, and the number is still growing. Production of monoclonal antibodies is a multistage, time-consuming and costly process. Growing demand for these molecules creates space for research focused on improvements in hybridoma technology. Lower costs, human labor, and time are important goals of these attempts. In this article, a brief review of current methods and their advances is given. PMID:27117113

  5. A homogeneous fluorescence-based method to measure antibody internalization in tumor cells.

    PubMed

    Gong, Haibiao; Urlacher, Teresa

    2015-01-15

    We have developed a simple fluorescence-based method to monitor antibody internalization. Panitumumab was dual-labeled with the fluorophore IRDye 800CW and quencher IRDye QC-1 to yield the biomolecular probe Pan800QC. The fluorescence of IRDye 800CW is quenched by IRDye QC-1 on the same intact antibody. After incubation with epidermal growth factor receptor (EGFR)-expressing cells, internalization of Pan800QC was detected by an increase in fluorescence signal due to enzymatic digestion of the antibody and separation of IRDye 800CW and IRDye QC-1. By optimizing reaction conditions, a signal-to-background ratio of 8.5 was obtained. This homogeneous assay can be applied in the characterization and screening of internalizing antibodies. PMID:25245185

  6. Methods of preparing and using single chain anti-tumor antibodies

    SciTech Connect

    Cheung, Nai-Kong; Guo, Hong-Fen

    2010-02-23

    This invention provides a method for identifying cells expressing a target single chain antibody (scFv) directed against a target antigen from a collection of cells that includes cells that do not express the target scFv, comprising the step of combining the collection of cells with an anti-idiotype directed to an antibody specific for the target antigen and detecting interaction, if any, of the anti-idiotype with the cells, wherein the occurrence of an interaction identifies the cell as one which expresses the target scFv. This invention also provides a method for making a single chain antibody (scFv) directed against an antigen, wherein the selection of clones is made based upon interaction of those clones with an appropriate anti-idiotype, and heretofore inaccessible scFv so made. This invention provides the above methods or any combination thereof. Finally, this invention provides various uses of these methods.

  7. A novel screening method to assess developability of antibody-like molecules

    PubMed Central

    Kohli, Neeraj; Jain, Nidhi; Geddie, Melissa L; Razlog, Maja; Xu, Lihui; Lugovskoy, Alexey A

    2015-01-01

    Monoclonal antibodies and antibody-like molecules represent a fast-growing class of bio-therapeutics that has rapidly transformed patient care in a variety of disease indications. The discovery of antibodies that bind to particular targets with high affinity is now a routine exercise and a variety of in vitro and in vivo techniques are available for this purpose. However, it is still challenging to identify antibodies that, in addition to having the desired biological effect, also express well, remain soluble at different pH levels, remain stable at high concentrations, can withstand high shear stress, and have minimal non-specific interactions. Many promising antibody programs have ultimately failed in development due to the problems associated with one of these factors. Here, we present a simple high-performance liquid chromatography (HPLC)-based screening method to assess these developability factors earlier in discovery process. This method is robust and requires only microgram quantities of proteins. Briefly, we show that for antibodies injected on a commercially available pre-packed Zenix HPLC column, the retention times are inversely related to their colloidal stability with antibodies prone to precipitation or aggregation retained longer on the column with broader peaks. By simply varying the salt content of running buffer, we were also able to estimate the nature of interactions between the antibodies and the column. We believe this approach should generally be applicable to assessment of the developability of other classes of bio-therapeutic molecules, and that the addition of this simple tool early in the discovery process will lead to selection of molecules with improved developability characteristics. PMID:25961854

  8. Lubrication approximation in completed double layer boundary element method

    NASA Astrophysics Data System (ADS)

    Nasseri, S.; Phan-Thien, N.; Fan, X.-J.

    This paper reports on the results of the numerical simulation of the motion of solid spherical particles in shear Stokes flows. Using the completed double layer boundary element method (CDLBEM) via distributed computing under Parallel Virtual Machine (PVM), the effective viscosity of suspension has been calculated for a finite number of spheres in a cubic array, or in a random configuration. In the simulation presented here, the short range interactions via lubrication forces are also taken into account, via the range completer in the formulation, whenever the gap between two neighbouring particles is closer than a critical gap. The results for particles in a simple cubic array agree with the results of Nunan and Keller (1984) and Stoksian Dynamics of Brady etal. (1988). To evaluate the lubrication forces between particles in a random configuration, a critical gap of 0.2 of particle's radius is suggested and the results are tested against the experimental data of Thomas (1965) and empirical equation of Krieger-Dougherty (Krieger, 1972). Finally, the quasi-steady trajectories are obtained for time-varying configuration of 125 particles.

  9. One-Step Conjugation Method for Site-Specific Antibody-Drug Conjugates through Reactive Cysteine-Engineered Antibodies.

    PubMed

    Shinmi, Daisuke; Taguchi, Eri; Iwano, Junko; Yamaguchi, Tsuyoshi; Masuda, Kazuhiro; Enokizono, Junichi; Shiraishi, Yasuhisa

    2016-05-18

    Engineered cysteine residues are particularly convenient for site-specific conjugation of antibody-drug conjugates (ADC), because no cell engineering and additives are required. Usually, unpaired cysteine residues form mixed disulfides during fermentation in Chinese hamster ovarian (CHO) cells; therefore, additional reduction and oxidization steps are required prior to conjugation. In this study, we prepared light chain (Lc)-Q124C variants in IgG and examined the conjugation efficiency. Intriguingly, Lc-Q124C exhibited high thiol reactivity and directly generated site-specific ADC without any pretreatment (named active thiol antibody: Actibody). Most of the cysteine-maleimide conjugates including Lc-Q124C showed retro-Michael reaction with cysteine 34 in albumin and were decomposed over time. In order to acquire resistance to a maleimide exchange reaction, the facile procedure for succinimide hydrolysis on anion exchange resin was employed. Hydrolyzed Lc-Q124C conjugate prepared with anion exchange procedure retained high stability in plasma. Recently, various stable linkage schemes for cysteine conjugation have been reported. The combination with direct conjugation by the use of Actibody and stable linker technology could enable the generation of stable site-specific ADC through a simple method. Actibody technology with Lc-Q124C at a less exposed position opens a new path for cysteine-based conjugation, and contributes to reducing entry barriers to the preparation and evaluation of ADC. PMID:27074832

  10. Development of Double Eastern Blotting for Major Licorice Components, Glycyrrhizin and Liquiritin for Chemical Quality Control of Licorice Using anti-Glycyrrhizin and anti-Liquiritin Monoclonal Antibodies.

    PubMed

    Fujii, Shunsuke; Morinaga, Osamu; Uto, Takuhiro; Nomura, Shuichi; Shoyama, Yukihiro

    2016-02-10

    Licorice is utilized in various food industries around the world for seasoning agents, confectioneries, drinks, and functional foods. Glycyrrhizin (GL) and liquiritin (Liq) are major quality control chemical markers of licorice that have multifunctional bioactivities. Chemical quality control of licorice is important because its component profiles change depending environmental factors (climate, soil condition, and water deficit) and differences between species. Double eastern blotting using anti-GL and anti-Liq monoclonal antibodies was developed for more convenient, rapid, and specific quality control analysis of GL and Liq, respectively. Moreover, double eastern blotting was applied to investigate the immunohistochemical distributions of GL and Liq in the root of fresh licorice; the localization of both components was then clarified visually. This double eastern blotting technique for GL and Liq may serve as a powerful approach for visually determining the chemical quality of licorice. PMID:26765784

  11. A comparative evaluation of six principal IgY antibody extraction methods.

    PubMed

    Ren, Hao; Yang, Wenjing; Thirumalai, Diraviyam; Zhang, Xiaoying; Schade, Rüdiger

    2016-03-01

    Egg yolk has been considered a promising source of antibodies. Our study was designed to compare six principal IgY extraction methods (water dilution, polyethylene glycol [PEG] precipitation, caprylic acid extraction, chloroform extraction, phenol extraction, and carrageenan extraction), and to assess their relative extraction efficiencies and the purity of the resulting antibodies. The results showed that the organic solvents (chloroform or phenol) minimised the lipid ratio in the egg yolk. The water dilution, PEG precipitation and caprylic acid extraction methods resulted in high yields, and antibodies purified with PEG and carrageenan exhibited high purity. Our results indicate that phenol extraction would be more suitable for preparing high concentrations of IgY for non-therapeutic usage, while the water dilution and carrageenan extraction methods would be more appropriate for use in the preparation of IgY for oral administration. PMID:27031600

  12. Novel method for the high-throughput production of phosphorylation site-specific monoclonal antibodies.

    PubMed

    Kurosawa, Nobuyuki; Wakata, Yuka; Inobe, Tomonao; Kitamura, Haruki; Yoshioka, Megumi; Matsuzawa, Shun; Kishi, Yoshihiro; Isobe, Masaharu

    2016-01-01

    Threonine phosphorylation accounts for 10% of all phosphorylation sites compared with 0.05% for tyrosine and 90% for serine. Although monoclonal antibody generation for phospho-serine and -tyrosine proteins is progressing, there has been limited success regarding the production of monoclonal antibodies against phospho-threonine proteins. We developed a novel strategy for generating phosphorylation site-specific monoclonal antibodies by cloning immunoglobulin genes from single plasma cells that were fixed, intracellularly stained with fluorescently labeled peptides and sorted without causing RNA degradation. Our high-throughput fluorescence activated cell sorting-based strategy, which targets abundant intracellular immunoglobulin as a tag for fluorescently labeled antigens, greatly increases the sensitivity and specificity of antigen-specific plasma cell isolation, enabling the high-efficiency production of monoclonal antibodies with desired antigen specificity. This approach yielded yet-undescribed guinea pig monoclonal antibodies against threonine 18-phosphorylated p53 and threonine 68-phosphorylated CHK2 with high affinity and specificity. Our method has the potential to allow the generation of monoclonal antibodies against a variety of phosphorylated proteins. PMID:27125496

  13. Novel method for the high-throughput production of phosphorylation site-specific monoclonal antibodies

    PubMed Central

    Kurosawa, Nobuyuki; Wakata, Yuka; Inobe, Tomonao; Kitamura, Haruki; Yoshioka, Megumi; Matsuzawa, Shun; Kishi, Yoshihiro; Isobe, Masaharu

    2016-01-01

    Threonine phosphorylation accounts for 10% of all phosphorylation sites compared with 0.05% for tyrosine and 90% for serine. Although monoclonal antibody generation for phospho-serine and -tyrosine proteins is progressing, there has been limited success regarding the production of monoclonal antibodies against phospho-threonine proteins. We developed a novel strategy for generating phosphorylation site-specific monoclonal antibodies by cloning immunoglobulin genes from single plasma cells that were fixed, intracellularly stained with fluorescently labeled peptides and sorted without causing RNA degradation. Our high-throughput fluorescence activated cell sorting-based strategy, which targets abundant intracellular immunoglobulin as a tag for fluorescently labeled antigens, greatly increases the sensitivity and specificity of antigen-specific plasma cell isolation, enabling the high-efficiency production of monoclonal antibodies with desired antigen specificity. This approach yielded yet-undescribed guinea pig monoclonal antibodies against threonine 18-phosphorylated p53 and threonine 68-phosphorylated CHK2 with high affinity and specificity. Our method has the potential to allow the generation of monoclonal antibodies against a variety of phosphorylated proteins. PMID:27125496

  14. An impedance spectroscopy method for the detection and evaluation of Babesia bovis antibodies in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An immunosensor method for diagnosis of Babesia bovis in cattle based on impedance measurement is presented in this study. The method probes the interaction between serum antibodies against B. bovis infected cattle and recombinant protein, RAP-1, with C-terminal obtained from a Portuguese B. bovis s...

  15. Methods for detection of West Nile virus antibodies in mosquito blood meals.

    PubMed

    Komar, Nicholas; Panella, Nicholas A; Young, Ginger R; Basile, Alison J

    2015-03-01

    We describe and compare 2 qualitative serologic techniques for detecting West Nile virus (WNV)-specific antibodies in mosquito blood meals. The techniques are the biotin microsphere immunoassay (b-MIA) and the inhibition platform of the VectorTest™ WNV antigen assay (VecTest-inhibition). To demonstrate the ability of these tests to detect WNV-neutralizing antibodies, we experimentally exposed feeding mosquitoes to blood containing 5 concentrations of 6B6C-1, a flavivirus-neutralizing monoclonal antibody. Antibody concentrations were quantified using the 90% plaque-reduction neutralization test (PRNT90). After 24 h of blood-meal digestion at 22.5°C, the threshold PRNT90 titer of detection was ≤18 for b-MIA and ≤50 for VecTest-inhibition. Both tests reliably detected antibodies in 3 of 3 blood meals that had been digested for up to 30 h, or were about 25% digested. The b-MIA was also applied to mosquitoes that had engorged on avian blood in Arizona following a WNV epidemic in 2010. There was no significant difference in the WNV antibody prevalence determined by b-MIA (52% of 71 avian blood meals) compared to the WNV-neutralizing antibody prevalence in birds determined by direct sampling (49% of 234 birds). VecTest-inhibition requires fewer resources and may be used in the field without a laboratory, but consumes the entire blood meal and relies on subjective interpretation of results. The b-MIA requires a laboratory and sophisticated equipment and reagents. Results for b-MIA are analyzed objectively and can be applied to mosquito blood meals with greater confidence than the VecTest-inhibition method and thus can contribute substantially to research and surveillance programs that would benefit from the detection of specific WNV antibodies in mosquito blood meals. PMID:25843170

  16. [Determination of anti-rabies antibodies by means of the indirect immunofluorescence method].

    PubMed

    Jiran, E; Závora, M

    1975-06-01

    The paper presents a description of the method of indirect immunofluorescence (IFRA test) for the demonstration of post-vaccination anti-rabies antibodies. The model of immune hamster serum was used to study the reproducibility of the test. The dynamics of antibody formation was quantitatively examined in rabbits immunized by the Flury-LEP virus. For brief information, the test was also used for the demonstration of post-vaccination antibodes against rabies in dogs. The method can be recommended as an additional and orientation test for the evaluation of the immunogenic properties of vaccines against rabies. PMID:810938

  17. Effects of Cationic Microbubble Carrying CD/TK Double Suicide Gene and αVβ3 Integrin Antibody in Human Hepatocellular Carcinoma HepG2 Cells

    PubMed Central

    Li, Jiale; Zhou, Ping; Li, Lan; Zhang, Yan; Shao, Yang; Tang, Li; Tian, Shuangming

    2016-01-01

    Objective Hepatocellular carcinoma (HCC), mostly derived from hepatitis or cirrhosisis, is one of the most common types of liver cancer. T-cell mediated immune response elicited by CD/TK double suicide gene has shown a substantial antitumor effect in HCC. Integrin αVβ3 over expresssion has been suggested to regulate the biology behavior of HCC. In this study, we investigated the strategy of incorporating CD/TK double suicide gene and anti-αVβ3 integrin monoclonal antibodies into cationic microbubbles (CMBsαvβ3), and evaluated its killing effect in HCC cells. Methods To improve the transfection efficiency of targeted CD/TK double suicide gene, we adopted cationic microbubbles (CMBs), a cationic delivery agent with enhanced DNA-carrying capacity. The ultrasound and high speed shearing method was used to prepare the non-targeting cationic microbubbles (CMBs). Using the biotin-avidin bridge method, αVβ3 integrin antibody was conjugated to CMBs, and CMBsαvβ3 was generated to specifically target to HepG2 cells. The morphology and physicochemical properties of the CMBsαvβ3 was detected by optical microscope and zeta detector. The conjugation of plasmid and the antibody in CMBsαvβ3 were examined by immunofluorescent microscopy and flow cytometry. The binding capacities of CMBsαvβ3 and CMBs to HCC HepG2 and normal L-02 cells were compared using rosette formation assay. To detect EGFP fluorescence and examine the transfection efficiencies of CMBsαvβ3 and CMBs in HCC cells, fluorescence microscope and contrast-enhanced sonography were adopted. mRNA and protein level of CD/TK gene were detected by RT-PCR and Western blot, respectively. To evaluate the anti-tumor effect of CMBsαvβ3, HCC cells with CMBsαvβ3 were exposed to 5-flurocytosine / ganciclovir (5-FC/GCV). Then, cell cycle distribution after treatment were detected by PI staining and flow cytometry. Apoptotic cells death were detected by optical microscope and assessed by MTT assay and TUNEL

  18. Phase 1 Randomized, Double-Blind, Placebo-Controlled Study of RG7667, an Anticytomegalovirus Combination Monoclonal Antibody Therapy, in Healthy Adults

    PubMed Central

    Ishida, Julie H.; Burgess, Tracy; Derby, Michael A.; Brown, Pearline A.; Maia, Mauricio; Deng, Rong; Emu, Brinda; Feierbach, Becket; Fouts, Ashley E.; Liao, X. Charlene

    2015-01-01

    Cytomegalovirus can cause debilitating and life-threatening disease in newborns infected in utero and immunocompromised individuals, including transplant recipients. RG7667 is a unique combination of two monoclonal antibodies that binds glycoprotein complexes on the surface of cytomegalovirus and inhibits its entry into host cells. A phase 1 first-in-human, randomized, double-blind, placebo-controlled, dose-escalation study of RG7667 given intravenously was conducted in 181 healthy adults. The study involved a single ascending dose stage (1, 3, 5, and 10 mg/kg each antibody; n = 21), a multiple ascending dose stage (5 and 10 mg/kg each antibody monthly for 3 doses; n = 10), and a multiple dose expansion stage (10 mg/kg each antibody monthly for 3 doses; n = 150). Subjects were followed for 85 to 141 days to evaluate safety, tolerability, pharmacokinetics, and immunogenicity. Most adverse events were mild, and the incidence of adverse events was similar among the RG7667 and placebo groups. RG7667 had dose-proportional pharmacokinetics in all three dosing stages, a mean terminal half-life of 20 to 30 days, and an overall pharmacokinetic profile consistent with that of a human monoclonal antibody that lacks endogenous host targets. The proportion of subjects developing an antitherapeutic antibody response was not higher in the RG7667 group than in the placebo group. In summary, single and multiple doses of RG7667 were found to be safe and well-tolerated in healthy adults and had a favorable pharmacokinetic and immunogenicity profile. This study supports further development of RG7667 as a therapy for the prevention and treatment of cytomegalovirus infection in susceptible populations. (This study has been registered at ClinicalTrials.gov under registration no. NCT01496755.) PMID:26055360

  19. Method to directly radiolabel antibodies for diagnostic imaging and therapy

    SciTech Connect

    Thakur, M.L.

    1991-04-30

    This patent describes a method for directly labeling proteins with radionuclides for use in diagnostic imaging and therapy. It comprises: the steps of incubating a protein-containing solution with a solution of sodium ascorbate; adding a required quantity of reduced radionuclide to the incubated protein-containing solution and incubating.

  20. High throughput peptide mapping method for analysis of site specific monoclonal antibody oxidation.

    PubMed

    Li, Xiaojuan; Xu, Wei; Wang, Yi; Zhao, Jia; Liu, Yan-Hui; Richardson, Daisy; Li, Huijuan; Shameem, Mohammed; Yang, Xiaoyu

    2016-08-19

    Oxidation of therapeutic monoclonal antibodies (mAbs) often occurs on surface exposed methionine and tryptophan residues during their production in cell culture, purification, and storage, and can potentially impact the binding to their targets. Characterization of site specific oxidation is critical for antibody quality control. Antibody oxidation is commonly determined by peptide mapping/LC-MS methods, which normally require a long (up to 24h) digestion step. The prolonged sample preparation procedure could result in oxidation artifacts of susceptible methionine and tryptophan residues. In this paper, we developed a rapid and simple UV based peptide mapping method that incorporates an 8-min trypsin in-solution digestion protocol for analysis of oxidation. This method is able to determine oxidation levels at specific residues of a mAb based on the peptide UV traces within <1h, from either TBHP treated or UV light stressed samples. This is the simplest and fastest method reported thus far for site specific oxidation analysis, and can be applied for routine or high throughput analysis of mAb oxidation during various stability and degradation studies. By using the UV trace, the method allows more accurate measurement than mass spectrometry and can be potentially implemented as a release assay. It has been successfully used to monitor antibody oxidation in real time stability studies. PMID:27432793

  1. Redoublement lexical, procede intensif (Lexical Doubling, Intensive Method).

    ERIC Educational Resources Information Center

    George, Kenneth E. M.

    1983-01-01

    An often-neglected aspect of daily language is syllable doubling or repetition, as in infant language ("nounou"), onomatopoeia ("ronron"), interjections or responses ("oui oui"), names ("Mimi"), or military slang ("coco" for "commandant"). The mechanisms and semantic functions of this phenomenon are outlined, drawing on examples from French…

  2. Histochemistry of nerve fibres double labelled with anti-TRPV2 antibodies and sensory nerve marker AM1-43 in the dental pulp of rat molars.

    PubMed

    Nishikawa, Sumio

    2008-09-01

    AM1-43 can label sensory nerve fibres and sensory neurons. Permeation of non-selective cation channels of the nerve cell membrane is suggested to be the mechanism responsible for labelling. To identify these channels, two candidates, TRPV1 and TRPV2 were examined by immunocytochemistry in the dental pulp and trigeminal ganglion of rats injected with AM1-43. A part of AM1-43-labelled nerve fibres was also positive for anti-TRPV2 antibody but negative for anti-TRPV1 antibody in the dental pulp. In the trigeminal ganglion, a part of the neuron showed both bright AM1-43 labelling and anti-TRPV2 immunolabelling, but neurons double labelled with AM1-43 and TRPV1 were rare. These results suggest that TRPV2 channels, but not TRPV1 channels, contribute to the fluorescent labelling of AM1-43 in the dental pulp. PMID:18405879

  3. Monoclonal antibodies to synthetic pyrethroids and method for detecting the same

    DOEpatents

    Stanker, L.H.; Vanderlaan, M.; Watkins, B.E.; Van Emon, J.M.; Bigbee, C.L.

    1992-04-28

    Methods are described for making specific monoclonal antibodies which may be used in a sensitive immunoassay for detection of synthetic pyrethroids in foods and environmental samples. Appropriate sample preparation and enzyme amplification of the immunoassay for this widely-used class of pesticides permits detection at low levels in laboratory and field tested samples. 6 figs.

  4. Monoclonal antibodies to synthetic pyrethroids and method for detecting the same

    DOEpatents

    Stanker, Larry H.; Vanderlaan, Martin; Watkins, Bruce E.; Van Emon, Jeanette M.; Bigbee, Carolyn L.

    1992-01-01

    Methods are described for making specific monoclonal antibodies which may be used in a sensitive immunoassay for detection of synthetic pyrethroids in foods and environmental samples. Appropriate sample preparation and enzyme amplification of the immunoassay for this widely-used class of pesticides permits detection at low levels in laboratory and field tested samples.

  5. An Improved Method for Estimating Antibody Titers in Microneutralization Assay Using Green Fluorescent Protein

    PubMed Central

    Yang, Hongmei; Baker, Steven F.; González, Mario E.; Topham, David J.; Martínez-Sobrido, Luis; Zand, Martin; Holden-Wiltse, Jeanne; Wu, Hulin

    2015-01-01

    Viruses that express reporter genes upon infection have been recently used to evaluate neutralizing antibody responses, where a lack of reporter expression indicates specific virus inhibition. The traditional model-based methods using standard outcome of percent neutralization could be applied to the data from the assays to estimate antibody titers. However, the data produced is sometimes irregular, which can yield meaningless outcomes of percent neutralization that do not fit the typical curves for immunoassays, making automated or semi-high throughput antibody titer estimation unreliable. We developed a type of new outcomes model, which is biologically meaningful and fits typical immunoassay curves well. Our simulation study indicates that the new response approach outperforms the traditional response approach regardless of the data variability. The proposed new response approach can be used in similar assays for other disease models. PMID:26010892

  6. Method to conjugate polysaccharide antigens to surfaces for the detection of antibodies.

    PubMed

    Boas, Ulrik; Lind, Peter; Riber, Ulla

    2014-11-15

    A new generic method for the conjugation of lipopolysaccharide (LPS)-derived polysaccharide antigens from gram-negative bacteria has been developed using Salmonella as a model. After removal of lipid A from the LPS by mild acidolysis, the polysaccharide antigen was conjugated to polystyrene microbeads modified with N-alkyl hydroxylamine and N-alkyl-O-methyl hydroxylamine surface groups by incubation of antigen and beads for 16 h at 40 °C without the need for coupling agents. The efficiency of the new method was evaluated by flow cytometry in model samples and serum samples containing antibodies against Salmonella typhimurium and Salmonella dublin. The presented method was compared with a similar method for conjugation of Salmonella polysaccharide antigens to surfaces. Here, the new method showed higher antigen coupling efficiency by detecting low concentrations of antibodies. Furthermore, the polysaccharide-conjugated beads showed preserved bioactivity after 1 year of use. PMID:25076184

  7. Comparative of three methods (ELIZA, MAIPA and flow cytometry) to determine anti-platelet antibody in children with ITP.

    PubMed

    Hamidpour, Mohsen; Khalili, Ghader; Tajic, Nader; Shamsian, Bi Bi Shahin; Hamidpour, Rafie

    2014-01-01

    Immune (idiopathic) thrombocytopenic purpurea (ITP) is an autoimmune disease characterized by the increased anti-platelet antibodies in the patient's sera and decreased platelets in the blood circulation. This study has determined and characterized the antiplatelet glycoproteins in children with ITP. Thirty eight children, who were hospitalized with clinical signs of ITP in Mofid Children Hospital (Tehran, Iran) during 18 months, went under our clinical studies in a research project. ELISA, Flow cytometry and MAIPA (Monoclonal Antibody Immobilization of Platelet Antigens) methods were employed to determine serum anti-platelet antibodies level. The anti-platelet antibodies level above mean + 3SD of control group was assumed as positive. The platelet counts ranged between 2 × 10(9)/L and 100 × 10(9)/L. Among the patients 63.5% of them were anti-platelet antibodies positive with ELISA method. Results of platelet lysate method showed that 51.7% of patients had antibodies against platelet antigens. Antibody against platelet GPIIb/IIIa, GPIb/IX and GPIa/IIa using MAIPA method were 48%, 54% and 25% respectively. In flow cytometry 62% of patients showed anti-platelet antibodies. The comparison of three methods shows that since MAIPA is the specific method for the detection of very small amount of antibody against glycoprotein antigens, it has the advantage of differentiating between immune and non-immune thrombocytopenia. PMID:25755908

  8. Comparative of three methods (ELIZA, MAIPA and flow cytometry) to determine anti-platelet antibody in children with ITP

    PubMed Central

    Hamidpour, Mohsen; Khalili, Ghader; Tajic, Nader; Shamsian, Bi Bi Shahin; Hamidpour, Rafie

    2014-01-01

    Immune (idiopathic) thrombocytopenic purpurea (ITP) is an autoimmune disease characterized by the increased anti-platelet antibodies in the patient’s sera and decreased platelets in the blood circulation. This study has determined and characterized the antiplatelet glycoproteins in children with ITP. Thirty eight children, who were hospitalized with clinical signs of ITP in Mofid Children Hospital (Tehran, Iran) during 18 months, went under our clinical studies in a research project. ELISA, Flow cytometry and MAIPA (Monoclonal Antibody Immobilization of Platelet Antigens) methods were employed to determine serum anti-platelet antibodies level. The anti-platelet antibodies level above mean + 3SD of control group was assumed as positive. The platelet counts ranged between 2 × 109/L and 100 × 109/L. Among the patients 63.5% of them were anti-platelet antibodies positive with ELISA method. Results of platelet lysate method showed that 51.7% of patients had antibodies against platelet antigens. Antibody against platelet GPIIb/IIIa, GPIb/IX and GPIa/IIa using MAIPA method were 48%, 54% and 25% respectively. In flow cytometry 62% of patients showed anti-platelet antibodies. The comparison of three methods shows that since MAIPA is the specific method for the detection of very small amount of antibody against glycoprotein antigens, it has the advantage of differentiating between immune and non-immune thrombocytopenia. PMID:25755908

  9. Double Cross-Validation in Multiple Regression: A Method of Estimating the Stability of Results.

    ERIC Educational Resources Information Center

    Rowell, R. Kevin

    In multiple regression analysis, where resulting predictive equation effectiveness is subject to shrinkage, it is especially important to evaluate result replicability. Double cross-validation is an empirical method by which an estimate of invariance or stability can be obtained from research data. A procedure for double cross-validation is…

  10. Interferometric Methods of Measuring Refractive Indices and Double-Refraction of Fibres.

    ERIC Educational Resources Information Center

    Hamza, A. A.; El-Kader, H. I. Abd

    1986-01-01

    Presents two methods used to measure the refractive indices and double-refraction of fibers. Experiments are described, with one involving the use of Pluta microscope in the double-beam interference technique, the other employing the multiple-beam technique. Immersion liquids are discussed that can be used in the experiments. (TW)

  11. Preferential association of a functional variant in complement receptor 2 with antibodies to double-stranded DNA

    PubMed Central

    Zhao, Jian; Giles, Brendan M; Taylor, Rhonda L; Yette, Gabriel A; Lough, Kara M; Ng, Han Leng; Abraham, Lawrence J; Wu, Hui; Kelly, Jennifer A; Glenn, Stuart B; Adler, Adam J; Williams, Adrienne H; Comeau, Mary E; Ziegler, Julie T; Marion, Miranda; Alarcón-Riquelme, Marta E; Alarcón, Graciela S; Anaya, Juan-Manuel; Bae, Sang-Cheol; Kim, Dam; Lee, Hye-Soon; Criswell, Lindsey A; Freedman, Barry I; Gilkeson, Gary S; Guthridge, Joel M; Jacob, Chaim O; James, Judith A; Kamen, Diane L; Merrill, Joan T; Sivils, Kathy Moser; Niewold, Timothy B; Petri, Michelle A; Ramsey-Goldman, Rosalind; Reveille, John D; Scofield, R Hal; Stevens, Anne M; Vilá, Luis M; Vyse, Timothy J; Kaufman, Kenneth M; Harley, John B; Langefeld, Carl D; Gaffney, Patrick M; Brown, Elizabeth E; Edberg, Jeffrey C; Kimberly, Robert P; Ulgiati, Daniela; Tsao, Betty P; Boackle, Susan A

    2016-01-01

    Objectives Systemic lupus erythematosus (SLE; OMIM 152700) is characterised by the production of antibodies to nuclear antigens. We previously identified variants in complement receptor 2 (CR2/CD21) that were associated with decreased risk of SLE. This study aimed to identify the causal variant for this association. Methods Genotyped and imputed genetic variants spanning CR2 were assessed for association with SLE in 15 750 case-control subjects from four ancestral groups. Allele-specific functional effects of associated variants were determined using quantitative real-time PCR, quantitative flow cytometry, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-PCR. Results The strongest association signal was detected at rs1876453 in intron 1 of CR2 (pmeta=4.2×10−4, OR 0.85), specifically when subjects were stratified based on the presence of dsDNA autoantibodies (case-control pmeta=7.6×10−7, OR 0.71; case-only pmeta=1.9×10−4, OR 0.75). Although allele-specific effects on B cell CR2 mRNA or protein levels were not identified, levels of complement receptor 1 (CR1/CD35) mRNA and protein were significantly higher on B cells of subjects harbouring the minor allele (p=0.0248 and p=0.0006, respectively). The minor allele altered the formation of several DNA protein complexes by EMSA, including one containing CCCTC-binding factor (CTCF), an effect that was confirmed by ChIP-PCR. Conclusions These data suggest that rs1876453 in CR2 has long-range effects on gene regulation that decrease susceptibility to lupus. Since the minor allele at rs1876453 is preferentially associated with reduced risk of the highly specific dsDNA autoantibodies that are present in preclinical, active and severe lupus, understanding its mechanisms will have important therapeutic implications. PMID:25180293

  12. Inverse scattering method and soliton double solution family for the general symplectic gravity model

    SciTech Connect

    Gao Yajun

    2008-08-15

    A previously established Hauser-Ernst-type extended double-complex linear system is slightly modified and used to develop an inverse scattering method for the stationary axisymmetric general symplectic gravity model. The reduction procedures in this inverse scattering method are found to be fairly simple, which makes the inverse scattering method applied fine and effective. As an application, a concrete family of soliton double solutions for the considered theory is obtained.

  13. [Study of nucleic acid structure by immunochemical methods. I. Antibodies specific for 6-sulfo-5,6-dihydro-4-methoxyaminopyrimidinone-2].

    PubMed

    Poverennyĭ, A M; Podgorodnichenko, V K; Monastyrskaia, G S; Bryskina, L E; Sverdlov, E D

    1978-01-01

    Immunization of animals with DNA modified by a mixture of bisulphite and O-methylhydroxylamine and methylated bovine serum albumin results in production of antibodies mainly reacting with modified DNA. Antibodies that react with denatured DNA were produced in minute quantity. It was shown that elicited antibodies possess a high specificity and have the ability to recognize only nucleotides with a double modification. The immune sera were fractionated by Sephadex G-200 column chromatography and the antibody activity was demonstrable in the 19S and 7S fractions. The attempts to induce synthesis of antibodies by injection of DNA modified by O-methylhydroxylamine failed. PMID:79979

  14. An improved method for covalently conjugating morpholino oligomers to antitumor antibodies.

    PubMed

    He, Jiang; Liu, Guozheng; Dou, Shuping; Gupta, Suresh; Rusckowski, Mary; Hnatowich, Donald

    2007-01-01

    Whether for conventional pretargeting, amplification pretargeting, or affinity enhancement pretargeting, it will be necessary to conjugate an antitumor antibody as the first injectate. This laboratory is investigating phosphorodiamidate morpholinos (MORFs) for pretargeting, and accordingly we are examining methods of attaching MORFs to antitumor antibodies that provide at least one group per molecule (gpm) without adversely influencing antibody properties. The aim of this investigation was to evaluate the commercial Hydralink for the conjugation of the anti-CEA MN14 antibody with an 18 mer amine-derivatized MORF. The conjugation was approached in both directions by first reacting MN14 with the NHS derivatives of 4-hydrozinonicotinate acetone hydrazone (SANH) or 4-formylbenzoate (SFB) and then combining with MORF that was previously reacted with SFB or SANH to yield MN14(SANH)-MORF and MN14(SFB)-MORF respectively. The storage stability, immunoreactive fraction, and the biodistribution in normal mice were compared for both conjugates. Thereafter, MN14(SANH)-MORF was used in a pretargeting study in tumored nude mice, and the results were compared to that obtained historically with MN14-MORF prepared by carbodiimide (EDC) coupling. Both new methods of conjugation provided between 1 and 2 gpm compared to 0.2 achieved previously by EDC. Furthermore, by repeat SE HPLC with and without CEA, both showed an unimpaired immunoreactive fraction. MN14(SANH)-MORF tolerated long-term storage best. More importantly, when labeled by hybridization with 99mTc-labeled complementary MORF (99mTc-cMORF), the biodistribution of MN14(SANH)-MORF was more favorable than that of MN14(SFB)-MORF in normal mice with lower liver (5.7 vs 9.4 %ID/g at 18 h) and spleen (3.5 vs 8.4 %ID/g) accumulations and higher blood levels (4.8 vs 3.4 %ID/g). Accordingly, only MN14(SANH)-MORF was used in a pretargeting study in tumored mice. When targeted with 99mTc-cMORF and at 2 days postinjection of antibody

  15. A method for double-labeling sputum cells for p53 and cytokeratin

    SciTech Connect

    Neft, R.E.; Tierney, L.A.; Belinsky, S.A.

    1995-12-01

    Molecular and immunological techniques may enhance the usefulness of sputum cytology as a screening tool for lung cancer. These techniques may also be useful in detecting and following the early progression of disease from metaplasia to dysplasia, carcinoma in situ, and finally to invasive carcinoma. Longitudinal information on the evolution of these malignant changes in the respiratory epithelium can be gained by prospective study of populations at high risk for lung cancer. This work is significant because double-labeling of cells in sputum with p53 and cytokeratin antibodies facilitates rapid screening of p53 positive neoplastic and preneoplastic lung cells by brightfield and fluorescence microscopy.

  16. MTF measurement of IRFPA based on double-knife edge scanning method

    NASA Astrophysics Data System (ADS)

    Ying, Cheng-ping; Wu, Bin; Wang, Heng-fei; Shi, Xue-shun; Liu, Hong-yuan

    2013-09-01

    Modulation transfer function (MTF) is one of the most important parameters of infrared focal plane array (IRFPA). A double-knife edge scanning method is proposed for MTF measurement of IRFPA. In this method, a double-knife edge was used as a target, and the IRFPA under test was positioned in the focal plane of the imaging optical system by a 3-axis translation stage. With an IRFPA data acquisition system, the image of the double-knife edge was restored. By scanning in the direction orthogonal to the double-knife edge image, edge spread function (ESF) curve of each pixel swept across the knife-edge image was obtained. MTF could be calculated from the subsequent fitting, differential and Fourier transformation procedures. With double-knife edge scanning, two ESF curves of double-knife edge were obtained simultaneously, and symmetry of the two ESF curves could be used to evaluate the verticality between photosensitive surface of IRFPA and optical axis of the double-knife edge imaging system. In addition, this method can be used to judge the existing of interference from outside such as vibration, stray light and electrical noise. A measurement facility for IRFPA's MTF based on double-knife edge scanning method was also established in this study. The facility is composed of double-knife edge imaging optical system, 3-axis translation stage and data acquisition system, et al. As the kernel of the facility, the double-knife edge imaging optical system mainly comprises two symmetrical parabolic mirrors coating with reflective material, and the magnification of the optical system is 1 with an operation wavelength range of (1˜14) μm.

  17. Detection of Aspergillus-specific antibodies by agar gel double immunodiffusion and IgG ELISA in feline upper respiratory tract aspergillosis.

    PubMed

    Barrs, V R; Ujvari, B; Dhand, N K; Peters, I R; Talbot, J; Johnson, L R; Billen, F; Martin, P; Beatty, J A; Belov, K

    2015-03-01

    Feline upper respiratory tract aspergillosis (URTA) is an emerging infectious disease. The aims of this study were: (1) to assess the diagnostic value of detection of Aspergillus-specific antibodies using an agar gel double immunodiffusion (AGID) assay and an indirect immunoglobulin G (IgG) ELISA; and (2) to determine if an aspergillin derived from mycelia of Aspergillus fumigatus, Aspergillus niger and Aspergillus flavus can be used to detect serum antibodies against cryptic Aspergillus spp. in Aspergillus section Fumigati. Sera from cats with URTA (group 1: n = 21) and two control groups (group 2: cats with other upper respiratory tract diseases, n = 25; group 3: healthy cats and cats with non-respiratory, non-fungal illness, n = 84) were tested. Isolates from cats with URTA comprised A. fumigatus (n = 5), A. flavus (n = 1) and four cryptic species: Aspergillus felis (n = 12), Aspergillus thermomutatus (Neosartorya pseudofischeri, n = 1), Aspergillus lentulus (n = 1) and Aspergillus udagawae (n = 1). Brachycephalic purebred cats were significantly more likely to develop URTA than other breeds (P = 0.013). The sensitivity (Se) of the AGID was 43% and the specificity (Sp) was 100%. At a cut-off value of 6 ELISA units/mL, the Se of the IgG ELISA was 95.2% and the Sp was 92% and 92.9% for groups 2 and 3 cats, respectively. Aspergillus-specific antibodies against all four cryptic species were detected in one or both assays. Assay Se was not associated with species identity. Detection of Aspergillus-specific antibodies by IgG ELISA has high Se and Sp for diagnosis of feline URTA. PMID:25634077

  18. MALDI immunoscreening (MiSCREEN): a method for selection of anti-peptide monoclonal antibodies for use in immunoproteomics.

    PubMed

    Razavi, Morteza; Pope, Matthew E; Soste, Martin V; Eyford, Brett A; Jackson, Angela M; Anderson, N Leigh; Pearson, Terry W

    2011-02-01

    A scalable method for screening and selection of peptide-specific monoclonal antibodies (mAbs) is described. To identify high affinity anti-peptide mAbs in hybridoma supernatants, antibodies were captured by magnetic affinity beads followed by binding of specific peptides from solution. After timed washing steps, the remaining bound peptides were eluted from the beads and detected by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). This allowed measurement of monovalent interactions of peptides with single antigen binding sites on the antibodies, thus reflecting antibody affinity rather than avidity. Antibodies that were able to bind target peptides from solution phase and retain them during washing for a minimum of 10 min were identified by the strength of the appropriate m/z peptide MS signals obtained. This wash time reflects the minimum peptide dissociation time required for use of these antibodies in several current immuno-mass spectrometry assays. Kinetic analysis of antibody-peptide binding by surface plasmon resonance (SPR) showed that the selected antibodies were of high affinity and, most importantly, had low dissociation constants. This method, called MALDI immunoscreening (MiSCREEN), thus enables rapid screening and selection of high affinity anti-peptide antibodies that are useful for a variety of immunoproteomics applications. To demonstrate their functional utility in immuno-mass spectrometry assays, we used the selected, purified RabMAbs to enrich natural (tryptic) peptides from digested human plasma. PMID:21078325

  19. Conjugation behaviours of CdTe quantum dots and antibody by a novel immunochromatographic method.

    PubMed

    Wang, Y; Bai, Y; Wei, X

    2011-03-01

    Three water-soluble CdTe quantum dots (QDs) (green-emitting, yellow-emitting and red-emitting) were synthesised for different refluxing time with 3-mercaptopropionic acid (MPA) as stabiliser. Then the red-emitting CdTe QDs and mouse immunoglobulin G (IgG) were taken as the representative to study the conjugation behaviour of QDs and antibody by a novel immunochromatographic method. After comparing with several methods, that is, direct conjugation, 1-ethyl-3(3-dimethylaminopropyl) carbodiimides hydrochloride (EDC)-mediated conjugation, N-hydroxysuccinimide (NHS)-mediated conjugation, EDC/NHS-mediated conjugation by immunochromatographic strips, EDC and NHS were selected together as coupling agents to conjugate QDs with antibody efficiently. Finally, the K562 leukaemia cells were incubated with the EDC/NHS-mediated conjugates to evaluate the performance in practical application, and the result from fluorescence images showed that it was successfully applied to label cells. The immunochromatographic strip was a superior method to study the conjugation of the fluorophore and antibody. PMID:21241157

  20. Production of anti-SAG1 IgY antibody against Toxoplasma gondii parasites and evaluation of antibody activity by ELISA method.

    PubMed

    Cakir-Koc, Rabia

    2016-08-01

    Chicken egg yolk antibody, also known as immunoglobulin Y (IgY), is the predominant class of serum immunoglobulin in birds. IgY has many advantages over mammalian antibodies, such as enhanced immunogenicity conserved mammalian proteins exhibited in birds due to their phylogenetic distance, non-invasive rapid, and economical collection system. However, there are limited studies about IgY production against Toxoplasma, which is a worldwide veterinary and public health problem. In this study, the production of specific IgY antibodies against the surface antigen 1 (SAG1) protein of Toxoplasma gondii and the determination of antibody activity via the enzyme-linked immunosorbent assay (ELISA) method were conducted. According to ELISA, Western blot, and NanoDrop results, specific and higher amounts of IgY antibody against SAG1 were obtained with this study. Considering the advantages of IgY and importance of SAG1 for the diagnosis of toxoplasmosis, it is expected that anti-SAG1 IgY will play an increasing role and gain commercial value in research, diagnostics, and immunotherapy against toxoplasmosis in the future. PMID:27079459

  1. Double-antibody sandwich enzyme-linked immunosorbent assay for quantitation of endoglucanase I of Trichoderma reesei.

    PubMed Central

    Bühler, R

    1991-01-01

    A sensitive and specific enzyme-liked immunosorbent assay for endoglucanase I (EG-I) has been developed. The monoclonal antibody a-EG-I 2, directed against an epitope on the core part of the enzyme, was used to capture the antigen in microtiter plate wells. A second, polyclonal antibody against the enzyme was then used to detect and quantitate the bound antigen. The test was specific for EG-I; neither endoglucanase II nor cellobiohydrolase I or II interfered. As little as 20 pg of EG-I protein could be detected. The coefficients of variation were 3.8% within plates and 6% between plates for a diluted Trichoderma reesei culture supernatant that contained 31 ng of EG-I per ml. Binding of the antigen to the monoclonal antibody was pH dependent and restricted to values between pH 6.5 and 10.5 with a maximum around pH 9. Standard solutions of EG-I were very stable at concentrations as low as 5 ng/ml when prepared in buffer that contained 1% bovine serum albumin and that was stored at -20 degrees C. After 37 weeks the antigenicity was still 97%. With this test it was possible to monitor the production of EG-I in a cellulase-producing strain of T. reesei and to demonstrate the apparent absence of the enzyme in a strain with the eglI gene deleted. PMID:1781689

  2. The double exponential sinc collocation method for singular Sturm-Liouville problems

    NASA Astrophysics Data System (ADS)

    Gaudreau, P.; Slevinsky, R.; Safouhi, H.

    2016-04-01

    Sturm-Liouville problems are abundant in the numerical treatment of scientific and engineering problems. In the present contribution, we present an efficient and highly accurate method for computing eigenvalues of singular Sturm-Liouville boundary value problems. The proposed method uses the double exponential formula coupled with sinc collocation method. This method produces a symmetric positive-definite generalized eigenvalue system and has exponential convergence rate. Numerical examples are presented and comparisons with single exponential sinc collocation method clearly illustrate the advantage of using the double exponential formula.

  3. Disjoint combinations profiling (DCP): a new method for the prediction of antibody CDR conformation from sequence

    PubMed Central

    Nikoloudis, Dimitris; Pitts, Jim E.

    2014-01-01

    The accurate prediction of the conformation of Complementarity-Determining Regions (CDRs) is important in modelling antibodies for protein engineering applications. Specifically, the Canonical paradigm has proved successful in predicting the CDR conformation in antibody variable regions. It relies on canonical templates which detail allowed residues at key positions in the variable region framework or in the CDR itself for 5 of the 6 CDRs. While no templates have as yet been defined for the hypervariable CDR-H3, instead, reliable sequence rules have been devised for predicting the base of the CDR-H3 loop. Here a new method termed Disjoint Combinations Profiling (DCP) is presented, which contributes a considerable advance in the prediction of CDR conformations. This novel method is explained and compared with canonical templates and sequence rules in a 3-way blind prediction. DCP achieved 93% accuracy over 951 blind predictions and showed an improvement in cumulative accuracy compared to predictions with canonical templates or sequence rules. In addition to its overall improvement in prediction accuracy, it is suggested that DCP is open to better implementations in the future and that it can improve as more antibody structures are deposited in the databank. In contrast, it is argued that canonical templates and sequence rules may have reached their peak. PMID:25071985

  4. Disjoint combinations profiling (DCP): a new method for the prediction of antibody CDR conformation from sequence.

    PubMed

    Nikoloudis, Dimitris; Pitts, Jim E; Saldanha, José W

    2014-01-01

    The accurate prediction of the conformation of Complementarity-Determining Regions (CDRs) is important in modelling antibodies for protein engineering applications. Specifically, the Canonical paradigm has proved successful in predicting the CDR conformation in antibody variable regions. It relies on canonical templates which detail allowed residues at key positions in the variable region framework or in the CDR itself for 5 of the 6 CDRs. While no templates have as yet been defined for the hypervariable CDR-H3, instead, reliable sequence rules have been devised for predicting the base of the CDR-H3 loop. Here a new method termed Disjoint Combinations Profiling (DCP) is presented, which contributes a considerable advance in the prediction of CDR conformations. This novel method is explained and compared with canonical templates and sequence rules in a 3-way blind prediction. DCP achieved 93% accuracy over 951 blind predictions and showed an improvement in cumulative accuracy compared to predictions with canonical templates or sequence rules. In addition to its overall improvement in prediction accuracy, it is suggested that DCP is open to better implementations in the future and that it can improve as more antibody structures are deposited in the databank. In contrast, it is argued that canonical templates and sequence rules may have reached their peak. PMID:25071985

  5. POLYTENE CHROMOSOME SQUASH METHODS FOR STUDYING TRANSCRIPTION AND EPIGENETIC CHROMATIN MODIFICATION IN DROSOPHILA USING ANTIBODIES

    PubMed Central

    Johansen, Kristen M.; Cai, Weili; Deng, Huai; Bao, Xiaomin; Zhang, Weiguo; Girton, Jack; Johansen, Jørgen

    2009-01-01

    The giant polytene chromosomes from Drosophila third instar larval salivary glands provide an important model system for studying the architectural changes in chromatin morphology associated with the process of transcription initiation and elongation. Especially, analysis of the heat shock response has proved useful in correlating chromatin structure remodeling with transcriptional activity. An important tool for such studies is the labeling of polytene chromosome squash preparations with antibodies to the enzymes, transcription factors, or histone modifications of interest. However, in any immunohistochemical experiment there will be advantages and disadvantages to different methods of fixation and sample preparation, the relative merits of which must be balanced. Here we provide detailed protocols for polytene chromosome squash preparation and discuss their relative pros and cons in terms of suitability for reliable antibody labeling and preservation of high resolution chromatin structure. PMID:19272452

  6. Single versus double testing of meat-juice samples for Salmonella antibodies, in the Danish pig-herd surveillance programme.

    PubMed

    Ekeroth, Lars; Alban, Lis; Feld, Niels

    2003-08-01

    In Denmark, a national serological surveillance-and-control programme for Salmonella in pigs has been in operation since 1995. The programme is based on the Danish mix-ELISA and uses double testing (two ELISA-wells used per sample) of meat-juice samples taken in relation to slaughter. All herds are classified monthly into one of the three levels; the classification is based on the percentage of positive serological results in the previous 3 months. In connection with evaluation of the programme in 2001, we investigated whether single testing (testing in one well only) could be expected to be sufficiently precise compared to double testing. Data from the year 2000 were used, and mathematical modelling. Single testing was simulated by randomised selection of one of the two results in the double testing. A slight increase in the prevalence of Salmonella-positive samples (1.02-1.09 times more through the four quarters of the year 2000) was found in the simulated single testing, as compared to the double testing. Around 0.5% of the herds would be allocated to another herd level in single testing-almost equal numbers one level up and one level down. No herd being seronegative in double testing would be allocated to levels 2 or 3 (herds with >40 or >70%, respectively, serological reactors) in single testing. The prevalence of "false-positive" diagnoses (positive in single testing and negative in double testing) and inversely defined "false-negative" diagnoses varied from 4.2 to 8.7% and from 3.2 to 4.5%, respectively, through the four quarters of the year 2000. The probability of allocating a herd to a wrong level due to sampling error was on the average 6.2 (varying from 1.66 to over 100) times higher than the probability of allocating a herd to a wrong level due to the test inaccuracy introduced by going from double to single testing. This is, however, an average; a herd with a true prevalence close to one of the level border cut-offs (40 and 70% weighted seroprevalence

  7. Detection of vibrio cholerae O1 by using cerium oxide nanowires - based immunosensor with different antibody immobilization methods

    NASA Astrophysics Data System (ADS)

    Tam, Phuong Dinh; Hoang, Nguyen Luong; Lan, Hoang; Vuong, Pham Hung; Anh, Ta Thi Nhat; Huy, Tran Quang; Thuy, Nguyen Thanh

    2016-05-01

    In this work, we evaluated the effects of different antibody immobilization strategies on the response of a CeO2-nanowires (NWs)-based immunosensor for Vibrio cholerae O1 detection. Accordingly, the changes in the electron-transfer resistance ( R et ) from before to after cells bind to an antibody-modified electrode prepared by using three different methods of antibody immobilization were determined. The values were 16.2%, 8.3%, and 6.65% for the method that utilized protein A, antibodies activated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS), and absorption, respectively. Cyclic voltammetry confirmed that the change in the current was highest for the immunosensors prepared using protein A (11%), followed by those prepared with EDC/NHS-activated antibodies (9%), and finally, those prepared through absorption (7.5%). The order of the antibody immobilization strategies in terms of resulting immunosensor detection limit and sensitivity was as follows order: absorption (3.2 × 103 CFU/mL; 45.1 Ω/CFU·mL-1) < EDC/NHS-activated antibody (1.0 × 103 CFU/mL; 50.6 Ω/CFU·mL-1) < protein A (1.0 × 102 CFU/mL; 65.8 Ω/CFU·mL-1). Thus, we confirmed that the protein A - mediated method showed significantly high cell binding efficiencies compared to the random immobilization method.

  8. Platelet antibody detection by flow cytometry: an effective method to evaluate and give transfusional support in platelet refractoriness

    PubMed Central

    Bub, Carolina Bonet; Martinelli, Beatriz Moraes; Avelino, Thayná Mendonça; Gonçalez, Ana Cláudia; Barjas-Castro, Maria de Lourdes; Castro, Vagner

    2013-01-01

    Background Immune platelet refractoriness is mainly caused by human leukocyte antigen antibodies (80-90% of cases) and, to a lesser extent, by human platelet antigen antibodies. Refractoriness can be diagnosed by laboratory tests and patients should receive compatible platelet transfusions. A fast, effective and low cost antibody-screening method which detects platelet human leukocyte/platelet antigen antibodies is essential in the management of immune platelet refractoriness. Objective The aim of this study was to evaluate the efficiency of the flow cytometry platelet immunofluorescence test to screen for immune platelet refractoriness. Methods A group of prospective hematologic patients with clinically suspected platelet refractoriness treated in a referral center in Campinas, SP during July 2006 and July 2011 was enrolled in this study. Platelet antibodies were screened using the flow cytometry platelet immunofluorescence test. Anti-human leukocyte antigen antibodies were detected by commercially available methods. The sensitivity, specificity and predictive values of the immunofluorescence test were determined taking into account that the majority of antiplatelet antibodies presented human leukocyte antigen specificity. Results Seventy-six samples from 32 female and 38 male patients with a median age of 43.5 years (range: 5-84 years) were analyzed. The sensitivity of the test was 86.11% and specificity 75.00% with a positive predictive value of 75.61% and a negative predictive value of 85.71%. The accuracy of the method was 80.26%. Conclusion This study shows that the flow cytometry platelet immunofluorescence test has a high correlation with the anti-human leukocyte antigen antibodies. Despite a few limitations, the method seems to be efficient, fast and feasible as the initial screening for platelet antibody detection and a useful tool to crossmatch platelets for the transfusional support of patients with immune platelet refractoriness. PMID:24106442

  9. The double-assignment method for the exponential chaotic tabu search in quadratic assignment problems

    NASA Astrophysics Data System (ADS)

    Shibata, Kazuaki; Horio, Yoshihiko; Aihara, Kazuyuki

    The quadratic assignment problem (QAP) is one of the NP-hard combinatorial optimization problems. An exponential chaotic tabu search using a 2-opt algorithm driven by chaotic neuro-dynamics has been proposed as one heuristic method for solving QAPs. In this paper we first propose a new local search, the double-assignment method, suitable for the exponential chaotic tabu search, which adopts features of the Lin-Kernighan algorithm. We then introduce chaotic neuro-dynamics into the double-assignment method to propose a novel exponential chaotic tabu search. We further improve the proposed exponential chaotic tabu search with the double-assignment method by enhancing the effect of chaotic neuro-dynamics.

  10. Assessing analytical methods to monitor isoAsp formation in monoclonal antibodies

    PubMed Central

    Eakin, Catherine M.; Miller, Amanda; Kerr, Jennifer; Kung, James; Wallace, Alison

    2014-01-01

    A ubiquitous post-translational modification observed in proteins is isomerization of aspartic acid to isoaspartic acid (isoAsp). This non-enzymatic post-translational modification occurs spontaneously in proteins and plays a role in aging, autoimmune response, cancer, neurodegeneration, and other diseases. Formation of isoAsp is also a significant issue for recombinant monoclonal antibody based protein therapeutics particularly when isomerization occurs in a complementarity-determining region due to potential impact to the clinical efficacy. Here, we present and compare three analytical methods to monitor and/or quantify isoAsp formation in a monoclonal antibody. The methods include two peptide map based technologies with quantitation from either UV integration or total ion peak areas, as well as an alternative approach using IdeS digestion to generate Fc/2 and Fab’2 regions, followed by hydrophobic interaction chromatography (HIC) to separate the population of Fab’2 containing an isoAsp. The level of isoAsp detected by the peptide map and the digested-HIC methods presented here show similar trends although sample throughput varies by method. PMID:24808864

  11. Simple Method To Prepare Oligonucleotide-Conjugated Antibodies and Its Application in Multiplex Protein Detection in Single Cells.

    PubMed

    Gong, Haibiao; Holcomb, Ilona; Ooi, Aik; Wang, Xiaohui; Majonis, Daniel; Unger, Marc A; Ramakrishnan, Ramesh

    2016-01-20

    The diversity of nucleic acid sequences enables genomics studies in a highly multiplexed format. Since multiplex protein detection is still a challenge, it would be useful to use genomics tools for this purpose. This can be accomplished by conjugating specific oligonucleotides to antibodies. Upon binding of the oligonucleotide-conjugated antibodies to their targets, the protein levels can be converted to oligonucleotide levels. In this report we describe a simple method for preparing oligonucleotide-conjugated antibodies and discuss this method's application in oligonucleotide extension reaction (OER) for multiplex protein detection. Conjugation is based on strain-promoted alkyne-azide cycloaddition (the Cu-free click reaction), in which the antibody is activated with a dibenzocyclooctyne (DBCO) moiety and subsequently linked covalently with an azide-modified oligonucleotide. In the functional test, the reaction conditions and purification processes were optimized to achieve maximum yield and best performance. The OER assay employs a pair of antibody binders (two antibodies, each conjugated with its own oligonucleotide) developed for each protein target. The two oligonucleotides contain unique six-base complementary regions at their 3' prime ends to allow annealing and extension by DNA synthesis enzymes to form a DNA template. Following preamplification, the DNA template is detected by qPCR. Distinct oligonucleotide sequences are assigned to different antibody binders to enable multiplex protein detection. When tested using recombinant proteins, some antibody binders, such as those specific to CSTB, MET, EpCAM, and CASP3, had dynamic ranges of 5-6 logs. The antibody binders were also used in a multiplexed format in OER assays, and the binders successfully detected their protein targets in cell lysates, and in single cells in combination with the C1 system. This click reaction-based antibody conjugation procedure is cost-effective, needs minimal hands-on time, and

  12. Extracorporeal adsorption therapy: A Method to improve targeted radiation delivered by radiometal-labeled monoclonal antibodies.

    SciTech Connect

    Nemecek, Eneida R.; Green, Damian J.; Fisher, Darrell R.; Pagal, John M.; Lin, Yukang; Gopal, A. K.; Durack, Lawrence D.; Rajendran, Joseph G.; Wilbur, D. S.; Nilsson, Rune; Sandberg, Bengt; Press, Oliver W.

    2008-04-01

    Many investigators have demonstrated the ability to treat hematologic malignancies with radiolabeled monoclonal antibodies targeting hematopoietic antigens such as anti-CD20 and anti-CD45. [1-5] Although the remission rates achieved with radioimmunotherapy (RIT) are relatively high, many patients subsequently relapse presumably due to suboptimal delivery of enough radiation to eradicate the malignancy. The dose-response of leukemia and lymphoma to radiation has been proven. Substantial amounts of radiation can be delivered by RIT if followed by hematopoietic cell transplantation to rescue the bone marrow from myeloablation.[ref] However, the maximum dose of RIT that can be used is still limited by toxicity to normal tissues affected by nonspecific delivery of radiation. Efforts to improve RIT focus on improving the therapeutic ratios of radiation in target versus non-target tissues by removing the fraction of radioisotope that fails to bind to target tissues and circulates freely in the bloodstream perfusing non-target tissues. Our group and others have explored several alternatives for removal of unbound circulating antibody. [refs] One such method, extracorporeal adsorption therapy (ECAT) consists of removing unbound antibody by a method similar to plasmapheresis after critical circulation time and distribution of antibody into target tissues have been achieved. Preclinical studies of ECAT in murine xenograft models demonstrated significant improvement in therapeutic ratios of radioactivity. Chen and colleagues demonstrated that a 2-hour ECAT procedure could remove 40 to 70% of the radioactivity from liver, lung and spleen. [ref] Although isotope concentration in the tumor was initially unaffected, a 50% decrease was noted approximately 36 hours after the procedure. This approach was also evaluated in a limited phase I pilot study of patients with refractory B-cell lymphoma. [ref] After radiographic confirmation of tumor localization of a test dose of anti-CD20

  13. Quantitative Detection of the Foot-And-Mouth Disease Virus Serotype O 146S Antigen for Vaccine Production Using a Double-Antibody Sandwich ELISA and Nonlinear Standard Curves.

    PubMed

    Feng, Xia; Ma, Jun-Wu; Sun, Shi-Qi; Guo, Hui-Chen; Yang, Ya-Min; Jin, Ye; Zhou, Guang-Qing; He, Ji-Jun; Guo, Jian-Hong; Qi, Shu-yun; Lin, Mi; Cai, Hu; Liu, Xiang-Tao

    2016-01-01

    The efficacy of an inactivated foot-and-mouth disease (FMD) vaccine is mainly dependent on the integrity of the foot-and-mouth disease virus (FMDV) particles. At present, the standard method to quantify the active component, the 146S antigen, of FMD vaccines is sucrose density gradient (SDG) analysis. However, this method is highly operator dependent and difficult to automate. In contrast, the enzyme-linked immunosorbent assay (ELISA) is a time-saving technique that provides greater simplicity and sensitivity. To establish a valid method to detect and quantify the 146S antigen of a serotype O FMD vaccine, a double-antibody sandwich (DAS) ELISA was compared with an SDG analysis. The DAS ELISA was highly correlated with the SDG method (R2 = 0.9215, P<0.01). In contrast to the SDG method, the DAS ELISA was rapid, robust, repeatable and highly sensitive, with a minimum quantification limit of 0.06 μg/mL. This method can be used to determine the effective antigen yields in inactivated vaccines and thus represents an alternative for assessing the potency of FMD vaccines in vitro. But it still needs to be prospectively validated by analyzing a new vaccine preparation and determining the proper protective dose followed by an in vivo vaccination-challenge study to confirm the ELISA findings. PMID:26930597

  14. Quantitative Detection of the Foot-And-Mouth Disease Virus Serotype O 146S Antigen for Vaccine Production Using a Double-Antibody Sandwich ELISA and Nonlinear Standard Curves

    PubMed Central

    Feng, Xia; Ma, Jun-Wu; Sun, Shi-Qi; Guo, Hui-Chen; Yang, Ya-Min; Jin, Ye; Zhou, Guang-Qing; He, Ji-Jun; Guo, Jian-Hong; Qi, Shu-yun; Lin, Mi; Cai, Hu; Liu, Xiang-Tao

    2016-01-01

    The efficacy of an inactivated foot-and-mouth disease (FMD) vaccine is mainly dependent on the integrity of the foot-and-mouth disease virus (FMDV) particles. At present, the standard method to quantify the active component, the 146S antigen, of FMD vaccines is sucrose density gradient (SDG) analysis. However, this method is highly operator dependent and difficult to automate. In contrast, the enzyme-linked immunosorbent assay (ELISA) is a time-saving technique that provides greater simplicity and sensitivity. To establish a valid method to detect and quantify the 146S antigen of a serotype O FMD vaccine, a double-antibody sandwich (DAS) ELISA was compared with an SDG analysis. The DAS ELISA was highly correlated with the SDG method (R2 = 0.9215, P<0.01). In contrast to the SDG method, the DAS ELISA was rapid, robust, repeatable and highly sensitive, with a minimum quantification limit of 0.06 μg/mL. This method can be used to determine the effective antigen yields in inactivated vaccines and thus represents an alternative for assessing the potency of FMD vaccines in vitro. But it still needs to be prospectively validated by analyzing a new vaccine preparation and determining the proper protective dose followed by an in vivo vaccination-challenge study to confirm the ELISA findings. PMID:26930597

  15. Double Force Compensation Method to Enhance the Performance of a Null Balance Force Sensor

    NASA Astrophysics Data System (ADS)

    Choi, In-Mook; Choi, Dong-June; Kim, Soo Hyun

    2002-06-01

    Microforce measurement is becoming more essential as precision industries such as biomedicine, precision chemistry, semiconductor manufacturing, and so forth develop. A null balance method has been introduced in order to improve on force measurement performances involving a loadcell. The null-balance type force sensor is analyzed and designed for the improvement of measurement performances. The measurement range and the resolution are dependent on the force generation capacity and the various error sources. These characteristics are estimated and verified according to the mechanical sensitivity and the force compensation sensitivity. Two different coil systems are designed and tested experimentally. Double force compensation is proposed in order to obtain a large range and high resolution. The measurement range of the large coil system and the resolution of the small one are fully realized by the double compensation method. After manufacturing, a range over 300 gf and resolution under ± 0.1 mgf were obtained by the double compensation method.

  16. Capillary isoelectric focusing method development and validation for investigation of recombinant therapeutic monoclonal antibody.

    PubMed

    Suba, Dávid; Urbányi, Zoltán; Salgó, András

    2015-10-10

    Capillary isoelectric focusing (cIEF) is a basic and highly accurate routine analytical tool to prove identity of protein drugs in quality control (QC) and release tests in biopharmaceutical industries. However there are some "out-of-the-box" applications commercially available which provide easy and rapid isoelectric focusing solutions for investigating monoclonal antibody drug proteins. However use of these kits in routine testings requires high costs. A capillary isoelectric focusing method was developed and validated for identification testing of monoclonal antibody drug products with isoelectric point between 7.0 and 9.0. A method was developed providing good pH gradient for internal calibration (R(2)>0.99) and good resolution between all of the isoform peaks (R=2), minimizing the time and complexity of sample preparation (no urea or salt used). The method is highly reproducible and it is suitable for validation and method transfer to any QC laboratories. Another advantage of the method is that it operates with commercially available chemicals which can be purchased from any suppliers. The interaction with capillary walls (avoid precipitation and adsorption as far as possible) was minimized and synthetic isoelectric small molecular markers were used instead of peptide or protein based markers. The developed method was validated according to the recent ICH guideline (Q2(R1)). Relative standard deviation results were below 0.2% for isoelectric points and below 4% according to the normalized migration times. The method is robust to buffer components with different lot numbers and neutral capillaries with different type of inner coatings. The fluoro-carbon coated column was chosen because of costs-effectivity aspects. PMID:26025812

  17. Double-layered target and identification method of individual target correlated with evaporation residues

    NASA Astrophysics Data System (ADS)

    Kaji, D.; Morimoto, K.

    2015-08-01

    A double-layered target system and an identification method (target ID) for individual targets mounted on a rotating wheel using correlation with evaporation residues were newly developed for the study of superheavy elements (SHE). The target system can be used in three modes: conventional single-layered mode, double-layered mode, and energy-degrader mode. The target ID method can be utilized for masking a target, measuring an excitation function without changing the beam energy from the accelerator, and searching for SHE nuclides using multiple targets during a single irradiation.

  18. Identification of discriminant proteins through antibody profiling, methods and apparatus for identifying an individual

    DOEpatents

    Thompson, Vicki S; Lacey, Jeffrey A; Gentillon, Cynthia A; Apel, William A

    2015-03-03

    A method for determining a plurality of proteins for discriminating and positively identifying an individual based from a biological sample. The method may include profiling a biological sample from a plurality of individuals against a protein array including a plurality of proteins. The protein array may include proteins attached to a support in a preselected pattern such that locations of the proteins are known. The biological sample may be contacted with the protein array such that a portion of antibodies in the biological sample reacts with and binds to the proteins forming immune complexes. A statistical analysis method, such as discriminant analysis, may be performed to determine discriminating proteins for distinguishing individuals. Proteins of interest may be used to form a protein array. Such a protein array may be used, for example, to compare a forensic sample from an unknown source with a sample from a known source.

  19. Identification of discriminant proteins through antibody profiling, methods and apparatus for identifying an individual

    DOEpatents

    Apel, William A.; Thompson, Vicki S; Lacey, Jeffrey A.; Gentillon, Cynthia A.

    2016-08-09

    A method for determining a plurality of proteins for discriminating and positively identifying an individual based from a biological sample. The method may include profiling a biological sample from a plurality of individuals against a protein array including a plurality of proteins. The protein array may include proteins attached to a support in a preselected pattern such that locations of the proteins are known. The biological sample may be contacted with the protein array such that a portion of antibodies in the biological sample reacts with and binds to the proteins forming immune complexes. A statistical analysis method, such as discriminant analysis, may be performed to determine discriminating proteins for distinguishing individuals. Proteins of interest may be used to form a protein array. Such a protein array may be used, for example, to compare a forensic sample from an unknown source with a sample from a known source.

  20. Method based on the double sideband technique for the dynamic tracking of micrometric particles

    NASA Astrophysics Data System (ADS)

    Ramirez, Claudio; Lizana, Angel; Iemmi, Claudio; Campos, Juan

    2016-06-01

    Digital holography (DH) methods are of interest in a large number of applications. Recently, the double sideband (DSB) technique was proposed, which is a DH based method that, by using double filtering, provides reconstructed images without distortions and is free of twin images by using an in-line configuration. In this work, we implement a method for the investigation of the mobility of particles based on the DSB technique. Particle holographic images obtained using the DSB method are processed with digital picture recognition methods, allowing us to accurately track the spatial position of particles. The dynamic nature of the method is achieved experimentally by using a spatial light modulator. The suitability of the proposed tracking method is validated by determining the trajectory and velocity described by glass microspheres in movement.

  1. Novel EGFR mutation specific antibodies for NSCLC: Immunohistochemistry as a possible screening method for EGFR mutations

    PubMed Central

    Kato, Yasufumi; Peled, Nir; Wynes, Murry W.; Yoshida, Koichi; Pardo, Marta; Mascaux, Celine; Ohira, Tatsuo; Tsuboi, Masahiro; Matsubayashi, Jun; Nagao, Toshitaka; Ikeda, Norihiko; Hirsch, Fred R.

    2010-01-01

    Background Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) predict better outcome to EGFR tyrosine kinase inhibitors (TKIs). The most common mutations are exon 19 deletions (most frequently E746-A750) and L858R point mutation in exon 21. Here, we evaluated the accuracy of novel EGFR mutation specific antibodies in a Japanese cohort with NSCLC and compared to direct DNA sequencing and clinical outcome. Materials and methods Immunohistochemistry (IHC) using antibodies specific for the E746-A750 and L858R mutations in EGFR was performed on tissue microarrays of tumors from 70 gefitinib treated NSCLC patients. Extracted DNA was sequenced for mutational analysis of EGFR exons 18 to 21. Results DNA sequencing showed EGFR mutations in 41 patients (58.6%), and exon 19 deletions in 18 patients (25.7%), 61% (11/18) had a deletion in the range of E746-A750) and 12 (17.1%) had exon 21 mutations (L858R). IHC showed, for the E746-A750 and L858R mutations, sensitivity (81.8% and 75%), specificity (100%, 96.6%), PPV (100%, 81.8%) and NPV (96.7%, 94.9%). Analysis for objective response rates (ORR) and survival were not correlated to IHC staining, although the combined staining showed non-significant trends towards better overall survival for patients with EGFR mutations. Conclusions The mutation specific IHC antibodies have high sensitivity and specificity for pre-defined EFGR mutations and may be suitable for screening for these pre-defined mutations. However, negative IHC results require further mutation analyses prior to excluding EGFR-targeted therapy. PMID:20697298

  2. A new hybrid double divisor ratio spectra method for the analysis of ternary mixtures

    NASA Astrophysics Data System (ADS)

    Youssef, Rasha M.; Maher, Hadir M.

    2008-10-01

    A new spectrophotometric method was developed for the simultaneous determination of ternary mixtures, without prior separation steps. This method is based on convolution of the double divisor ratio spectra, obtained by dividing the absorption spectrum of the ternary mixture by a standard spectrum of two of the three compounds in the mixture, using combined trigonometric Fourier functions. The magnitude of the Fourier function coefficients, at either maximum or minimum points, is related to the concentration of each drug in the mixture. The mathematical explanation of the procedure is illustrated. The method was applied for the assay of a model mixture consisting of isoniazid (ISN), rifampicin (RIF) and pyrazinamide (PYZ) in synthetic mixtures, commercial tablets and human urine samples. The developed method was compared with the double divisor ratio spectra derivative method (DDRD) and derivative ratio spectra-zero-crossing method (DRSZ). Linearity, validation, accuracy, precision, limits of detection, limits of quantitation, and other aspects of analytical validation are included in the text.

  3. Identification of antibody isotypes in biological fluids by means of micro-Raman spectroscopy and chemometric methods

    NASA Astrophysics Data System (ADS)

    Araujo-Andrade, C.; Pichardo-Molina, J. L.; Barbosa-Sabanero, G.; Frausto-Reyes, C.

    2008-02-01

    Clinical diagnosis of infections, generally are realized by serological methods, which identifies the antibodies presents in serum or tissue fluids of the patient. Antibodies are proteins present in our bodies that aid in the elimination of pathogens or antigens. Identification of antibodies isotypes is important because can help to predict when and whether patients will recover from infections and are commonly diagnosed by means of indirect methods such as serological test. In the other hand, the majority of these methods requires specific kits for the analysis, special sample preparation, chemical reagents, expensive equipment and require long time for getting results. In this work we show the feasibility to discriminate antibody isotypes in biological fluids like human colostrum by means of Raman spectroscopy and chemometrics. Spectra were obtained using an excitation wavelength of 514 nm over dried samples of human colostrum labeled previously as positives to specific IgG and IgM antibodies against Toxoplasma Gondii by means of ELISA test. Partial least square-discriminant analysis (PLS-DA) was used to discriminate among antibody isotypes by use second derivative of Raman spectra of colostrum samples.

  4. A method to confer Protein L binding ability to any antibody fragment

    PubMed Central

    Lakhrif, Zineb; Pugnière, Martine; Henriquet, Corinne; di Tommaso, Anne; Dimier-Poisson, Isabelle; Billiald, Philippe; Juste, Matthieu O.; Aubrey, Nicolas

    2016-01-01

    abstract Recombinant antibody single-chain variable fragments (scFv) are difficult to purify homogeneously from a protein complex mixture. The most effective, specific and fastest method of purification is an affinity chromatography on Protein L (PpL) matrix. This protein is a multi-domain bacterial surface protein that is able to interact with conformational patterns on kappa light chains. It mainly recognizes amino acid residues located at the VL FR1 and some residues in the variable and constant (CL) domain. Not all kappa chains are recognized, however, and the lack of CL can reduce the interaction. From a scFv composed of IGKV10-94 according to IMGT®, it is possible, with several mutations, to transfer the motif from the IGKV12-46 naturally recognized by the PpL, and, with the single mutation T8P, to confer PpL recognition with a higher affinity. A second mutation S24R greatly improves the affinity, in particular by modifying the dissociation rate (kd). The equilibrium dissociation constant (KD) was measured at 7.2 10-11 M by surface plasmon resonance. It was possible to confer PpL recognition to all kappa chains. This protein interaction can be modulated according to the characteristics of scFv (e.g., stability) and their use with conjugated PpL. This work could be extrapolated to recombinant monoclonal antibodies, and offers an alternative for protein A purification and detection. PMID:26683650

  5. A FORTRAN Program for Computing Refractive Index Using the Double Variation Method.

    ERIC Educational Resources Information Center

    Blanchard, Frank N.

    1984-01-01

    Describes a computer program which calculates a best estimate of refractive index and dispersion from a large number of observations using the double variation method of measuring refractive index along with Sellmeier constants of the immersion oils. Program listing with examples will be provided on written request to the author. (Author/JM)

  6. Cutting-edge mass spectrometry methods for the multi-level structural characterization of antibody-drug conjugates.

    PubMed

    Beck, Alain; Terral, Guillaume; Debaene, François; Wagner-Rousset, Elsa; Marcoux, Julien; Janin-Bussat, Marie-Claire; Colas, Olivier; Dorsselaer, Alain Van; Cianférani, Sarah

    2016-02-01

    Antibody drug conjugates (ADCs) are highly cytotoxic drugs covalently attached via conditionally stable linkers to monoclonal antibodies (mAbs) and are among the most promising next-generation empowered biologics for cancer treatment. ADCs are more complex than naked mAbs, as the heterogeneity of the conjugates adds to the inherent microvariability of the biomolecules. The development and optimization of ADCs rely on improving their analytical and bioanalytical characterization by assessing several critical quality attributes, namely the distribution and position of the drug, the amount of naked antibody, the average drug to antibody ratio, and the residual drug-linker and related product proportions. Here brentuximab vedotin (Adcetris®) and trastuzumab emtansine (Kadcyla®), the first and gold-standard hinge-cysteine and lysine drug conjugates, respectively, were chosen to develop new mass spectrometry (MS) methods and to improve multiple-level structural assessment protocols. PMID:26653789

  7. The bovine immune response to Brucella abortus IV. Studies with a double immunodiffusion test for antibody against A2.

    PubMed Central

    Stemshorn, B; Nielsen, K

    1981-01-01

    A double immunodiffusion test for precipitins against Brucella antigen A2 was developed and applied to a variety of samples. The A2 precipitins were produced by a heifer infected with B. abortus strain 2308, cattle vaccinated with killed B. melitensis strain H38 or live B. abortus strain 19 and by a dog infected with B. canis. Precipitins were also detected in the second International Standard for anti-Brucella abortus serum, in several anti-B. canis sera and at low levels in one anti-B. ovis serum tested. Antisera produced in calves against Yersinia enterocolitica serotype 0:9 had no anti-A2 activity despite titers greater than or equal to 1/1024 and greater than or equal to 1/80 in standard Brucella agglutination and CF tests, respectively. The test for A2 precipitins lacked specificity as weak reactions were obtained with five of 295 sera from brucellosis-free herds. This test was relatively insensitive, detecting precipitins in only 16 of 24 sera from infected cattle and 27 of 54 sera positive by complement fixation and enzyme labelled antiglobulin tests performed with whole cell and smooth lipopolysaccharide antigens, respectively. The A2 precipitins were detected in nine sera from five cattle, in two infected herds, which were negative by agglutination and complement fixation tests. Images Fig. 1. Fig. 2. PMID:6790144

  8. Mean centering of double divisor ratio spectra, a novel spectrophotometric method for analysis of ternary mixtures.

    PubMed

    Hassan, Said A; Elzanfaly, Eman S; Salem, Maissa Y; El-Zeany, Badr A

    2016-01-15

    A novel spectrophotometric method was developed for determination of ternary mixtures without previous separation, showing significant advantages over conventional methods. The new method is based on mean centering of double divisor ratio spectra. The mathematical explanation of the procedure is illustrated. The method was evaluated by determination of model ternary mixture and by the determination of Amlodipine (AML), Aliskiren (ALI) and Hydrochlorothiazide (HCT) in laboratory prepared mixtures and in a commercial pharmaceutical preparation. For proper presentation of the advantages and applicability of the new method, a comparative study was established between the new mean centering of double divisor ratio spectra (MCDD) and two similar methods used for analysis of ternary mixtures, namely mean centering (MC) and double divisor of ratio spectra-derivative spectrophotometry (DDRS-DS). The method was also compared with a reported one for analysis of the pharmaceutical preparation. The method was validated according to the ICH guidelines and accuracy, precision, repeatability and robustness were found to be within the acceptable limits. PMID:26298680

  9. Mean centering of double divisor ratio spectra, a novel spectrophotometric method for analysis of ternary mixtures

    NASA Astrophysics Data System (ADS)

    Hassan, Said A.; Elzanfaly, Eman S.; Salem, Maissa Y.; El-Zeany, Badr A.

    2016-01-01

    A novel spectrophotometric method was developed for determination of ternary mixtures without previous separation, showing significant advantages over conventional methods. The new method is based on mean centering of double divisor ratio spectra. The mathematical explanation of the procedure is illustrated. The method was evaluated by determination of model ternary mixture and by the determination of Amlodipine (AML), Aliskiren (ALI) and Hydrochlorothiazide (HCT) in laboratory prepared mixtures and in a commercial pharmaceutical preparation. For proper presentation of the advantages and applicability of the new method, a comparative study was established between the new mean centering of double divisor ratio spectra (MCDD) and two similar methods used for analysis of ternary mixtures, namely mean centering (MC) and double divisor of ratio spectra-derivative spectrophotometry (DDRS-DS). The method was also compared with a reported one for analysis of the pharmaceutical preparation. The method was validated according to the ICH guidelines and accuracy, precision, repeatability and robustness were found to be within the acceptable limits.

  10. Humanization of predicted T-cell epitopes reduces the immunogenicity of chimeric antibodies: new evidence supporting a simple method.

    PubMed

    Roque-Navarro, Lourdes; Mateo, Cristina; Lombardero, Josefa; Mustelier, Geraudis; Fernández, Alicia; Sosa, Katya; Morrison, Sherrie L; Pérez, Rolando

    2003-08-01

    Genetic engineering has provided several approaches to reduce immunogenicity of murine antibodies. We described previously a new method based on the humanization of the linear epitopes presented to T cells. In brief, potential immunogenic epitopes in the variable region were identified and subjected to point mutations to make them human and/or to modify amphipatic motifs. The resulting recombinant antibody retained its antigen binding affinity and was less immunogenic in monkeys than their murine or chimeric predecessors are. The present study provides two new examples of this T-cell epitope humanization approach: ior-t1A murine monoclonal antibody (mMAb), which recognizes the human-CD6 molecule, and ior-C5 mMAb, which recognizes a novel glycoprotein expressed on the surface of malignant colorectal cells. Seven amino acids were substituted in ior-C5 and eleven residues in ior-t1A, by the corresponding residues from the highest homologous human sequences. Surprisingly, the homology between re-shaped chimeric antibody variable region frameworks and human sequences was 80-90%. Experiments in monkeys showed that T1AhT and C5hT "detopes" antibodies were less immunogenic than their chimeric analogues while they retained 30-50% of antigen binding affinities. The proposed method might be of general applicability to reduce immunogenicity of chimeric antibodies with therapeutic potential. PMID:14511570

  11. Double Hypernuclei Experiment with Hybrid Emulsion Method at J-PARC

    NASA Astrophysics Data System (ADS)

    Ekawa, Hiroyuki

    Double hypernuclei are important probes to study the system with strangeness S = -2. Several emulsion experiments had been performed to search for them. We are planning a new experiment to search for double hypernuclei at the K1.8 beam line in the Hadron Experimental Facility (J-PARC E07 experiment). Ξ- tracks in the emulsion plates and SSD will be automatically connected by a hybrid method. The estimated Ξ- stopped statistics is 10 times as high as that of the KEK E373 experiment. Discoveries of 10 new double hypernuclear species are expected, which enable us to discuss binding energy in terms of mass number dependence. On the other hand, we will also observe X rays from Ξ- atoms with a germanium detector array installed close to theemulsion plates by tagging Ξ- stopped events. This will be the first measurement to give information on the Ξ- potential at the nuclear surface region.

  12. An analytical method for analyzing symmetry-breaking bifurcation and period-doubling bifurcation

    NASA Astrophysics Data System (ADS)

    Zou, Keguan; Nagarajaiah, Satish

    2015-05-01

    A new modification of homotopy analysis method (HAM) is proposed in this paper. The auxiliary differential operator is specifically chosen so that more than one secular term must be eliminated. The proposed method can capture asymmetric and period-2 solutions with satisfactory accuracy and hence can be used to predict symmetry-breaking and period-doubling bifurcation points. The variation of accuracy is investigated when different number of frequencies are considered.

  13. Applying a kinetic method to an indirect ELISA measuring Ostertagia ostertagi antibodies in milk

    PubMed Central

    Vanderstichel, Raphaël; Dohoo, Ian; Markham, Fred

    2015-01-01

    Indirect enzyme-linked immunosorbent assays (ELISAs) are frequently run as endpoint ELISAs (e-ELISAs). However, kinetic ELISAs (k-ELISAs) have certain advantages over e-ELISAs. The objective of this study was to understand the relationship between e-ELISA and k-ELISA results. Specifically, to determine whether it was possible to run both k-ELISA and e-ELISA on the same plate and establish an appropriate time interval for k-ELISA measurements. A normalization method for k-ELISA slopes (slope ratio) is proposed. Using an indirect e-ELISA test measuring antibodies against Ostertagia ostertagi in milk from dairy cattle, we found that running a k-ELISA had no effect on optical density ratio results of an e-ELISA on the same plate, and that agreement was very strong at 10, 15, and 28 min, allowing for a reduction in the total processing time for ELISA tests. PMID:26130849

  14. Simple, rapid /sup 125/I-labeled cyclosporine double antibody/polyethylene glycol radioimmunoassay used in a pediatric cardiac transplant program

    SciTech Connect

    Berk, L.S.; Webb, G.; Imperio, N.C.; Nehlsen-Cannarella, S.L.; Eby, W.C.

    1986-01-01

    We modified the Sandoz cyclosporine radioimmunoassay because of our need for frequent clinical monitoring of cyclosporine drug levels in allo- and xenograft pediatric cardiac transplant patients. With application of a commercially available (/sup 125/I)cyclosporine label in place of (/sup 3/H)cyclosporine and a second antibody/polyethylene glycol (PEG) method of separation in place of charcoal separation, we simplified and enhanced the speed and precision of assay performance. Studies of 140 whole blood samples comparing this new method to the (/sup 3/H)cyclosporine radioimmunoassay (RIA) method of Berk and colleagues yielded a coefficient of correlation of 0.96 (p less than 0.00001) with means of 626 and 667 ng/ml for (/sup 3/H)RIA and (/sup 125/I)RIA, respectively, and a regression equation of y = 28 + 1.02x. The major advantages are that total assay time is reduced to approximately 1 h; (/sup 125/I)cyclosporine label is used, avoiding the problems associated with liquid scintillation counting; and precision is enhanced by separating bound and free fractions with second antibody/PEG. These modifications should provide for greater ease of assay performance and improved clinical utility of cyclosporine monitoring not only in the pediatric but also in the adult transplant patient.

  15. Complementary MS Methods Assist Conformational Characterization of Antibodies with Altered S-S Bonding Networks

    NASA Astrophysics Data System (ADS)

    Jones, Lisa M.; Zhang, Hao; Cui, Weidong; Kumar, Sandeep; Sperry, Justin B.; Carroll, James A.; Gross, Michael L.

    2013-06-01

    As therapeutic monoclonal antibodies (mAbs) become a major focus in biotechnology and a source of the next-generation drugs, new analytical methods or combination methods are needed for monitoring changes in higher order structure and effects of post-translational modifications. The complexity of these molecules and their vulnerability to structural change provide a serious challenge. We describe here the use of complementary mass spectrometry methods that not only characterize mutant mAbs but also may provide a general framework for characterizing higher order structure of other protein therapeutics and biosimilars. To frame the challenge, we selected members of the IgG2 subclass that have distinct disulfide isomeric structures as a model to evaluate an overall approach that uses ion mobility, top-down MS sequencing, and protein footprinting in the form of fast photochemical oxidation of proteins (FPOP). These three methods are rapid, sensitive, respond to subtle changes in conformation of Cys → Ser mutants of an IgG2, each representing a single disulfide isoform, and may be used in series to probe higher order structure. The outcome suggests that this approach of using various methods in combination can assist the development and quality control of protein therapeutics.

  16. Complementary MS Methods Assist Conformational Characterization of Antibodies with Altered S–S Bonding Networks

    PubMed Central

    Jones, Lisa M.; Zhang, Hao; Cui, Weidong; Kumar, Sandeep; Sperry, Justin B.; Carroll, James A.; Gross, Michael L.

    2013-01-01

    As therapeutic monoclonal antibodies (mAbs) become a major focus in biotechnology and a source of the next-generation drugs, new analytical methods or combination methods are needed for monitoring changes in higher order structure and effects of post-translational modifications. The complexity of these molecules and their vulnerability to structural change provide a serious challenge. We describe here the use of complementary mass spectrometry methods that not only characterize mutant mAbs but also may provide a general framework for characterizing higher order structure of other protein therapeutics and biosimilars. To frame the challenge, we selected members of the IgG2 subclass that have distinct disulfide isomeric structures as a model to evaluate an overall approach that uses ion mobility, top-down MS sequencing, and protein footprinting in the form of fast photochemical oxidation of proteins (FPOP). These three methods are rapid, sensitive, respond to subtle changes in conformation of Cys→Ser mutants of an IgG2, each representing a single disulfide isoform, and may be used in series to probe higher order structure. The outcome suggests that this approach of using various methods in combination can assist the development and quality control of protein therapeutics. PMID:23483515

  17. Investigation of an innovative method for DC flow suppression of double-inlet pulse tube coolers

    NASA Astrophysics Data System (ADS)

    Hu, J. Y.; Luo, E. C.; Wu, Z. H.; Dai, W.; Zhu, S. L.

    2007-05-01

    The use of double-inlet mode in the pulse tube cooler opens up a possibility of DC flow circulating around the regenerator and the pulse tube. The DC flow sometimes deteriorates the performance of the cryocooler because such a steady flow adds an unwanted thermal load to the cold heat exchanger. It seems that this problem is still not well solved although a lot of effort has been made. Here we introduce a membrane-barrier method for DC flow suppression in double-inlet pulse tube coolers. An elastic membrane is installed between the pulse tube cooler inlet and the double-inlet valve to break the closed-loop flow path of DC flow. The membrane is acoustically transparent, but would block the DC flow completely. Thus the DC flow is thoroughly suppressed and the merit of double-inlet mode is remained. With this method, a temperature reduction of tens of Kelvin was obtained in our single-stage pulse tube cooler and the lowest temperature reached 29.8 K.

  18. Application of magnetic printing method to hard-disk media with double recording layers

    NASA Astrophysics Data System (ADS)

    Ono, Takuya; Kuboki, Yoshiyuki; Ajishi, Yoshifumi; Saito, Akira

    2003-05-01

    The magnetic printing method, which can duplicate soft magnetic patterns containing digital information such as servosignals formed on a master disk onto recording media, enables signals to be written to hard-disk media having high coercivities above 6000 Oe. We propose the application of the magnetic printing method to a hard-disk medium having double recording layers, one layer of which has high coercivity and is to be printed with digital information. This double recording layer medium is a hard-disk medium that has a magnetic read-only-memory (MROM) layer. In this study, we demonstrated a method for printing to this medium, which has MROM, and discussed the magnetic properties and recording performances of this medium.

  19. Development of electrochemical immunosensors based on different serum antibody immobilization methods for detection of Japanese encephalitis virus

    NASA Astrophysics Data System (ADS)

    Tran, Quang Huy; Hanh Nguyen, Thi Hong; Mai, Anh Tuan; Thuy Nguyen, Thi; Khue Vu, Quang; Nga Phan, Thi

    2012-03-01

    This paper describes the development of electrochemical immunosensors based on human serum antibodies with different immobilization methods for detection of Japanese encephalitis virus (JEV). Human serum containing anti-JEV antibodies was used to immobilize onto the surface of silanized interdigitated electrodes by four methods: direct adsorption (APTES-serum), covalent binding with a cross linker of glutaraldehyde (APTES-GA-serum), covalent binding with a cross linker of glutaraldehyde combined with anti-human IgG (APTES-GA-anti-HIgG-serum) and covalent binding with a cross linker of glutaraldehyde combined with a bioaffinity of protein A (APTES-GA-PrA-serum). Atomic force microscopy was used to verify surface characteristics of the interdigitated electrodes before and after treatment with serum antibodies. The output signal of the immunosensors was measured by the change of conductivity resulting from the specific binding of JEV antigens and serum antibodies immobilized on the electrodes, with the help of horseradish peroxidase (HRP)-labeled secondary antibody against JEV. The results showed that the APTES-GA-PrA-serum method provided the highest signal of the electrochemical immunosensor for detection of JEV antigens, with the linear range from 25 ng ml‑1 to 1 μg ml‑1, and the limit of detection was about 10 ng ml‑1. This study shows a potential development of novel electrochemical immunosensors applied for virus detection in clinical samples in case of possible outbreaks.

  20. A double filtering method for measuring the translational velocity of fluorescently stained cells

    SciTech Connect

    Yasokawa, Toshiki; Ishimaru, Ichirou; Kuriyama, Shigeki; Masaki, Tsutomu; Takegawa, Kaoru; Tanaka, Naotaka

    2007-09-24

    The authors propose a double filtering method to measure translational velocity for tracking fluorescently stained cells. This method employs two diffraction gratings installed in the infinity space through which the parallel pencil beam of the fluorescence passes. With this method, the change in light intensity whose period is proportional to the translational velocity of the sample can be obtained at the imaging surface. By using a sample that has a random distribution of fluorescence intensity, the authors verified that translational velocity measurements could be achieved using the proposed method.

  1. Chromatography-based methods for determining molar extinction coefficients of cytotoxic payload drugs and drug antibody ratios of antibody drug conjugates.

    PubMed

    Wang, Chunlei; Chen, Sike; Caceres-Cortes, Janet; Huang, Richard Y-C; Tymiak, Adrienne A; Zhang, Yingru

    2016-07-15

    UV spectrophotometry is widely used to determine the molar extinction coefficients (MECs) of cytotoxic drugs as well as the drug antibody ratios (DARs) of antibody drug conjugates (ADCs). However, the unknown purity of a drug due to interfering impurities can lead to erroneous MECs and DARs. Hence, reliable methods to accurately determine purity and the MECs of drugs with limited quantity is urgently needed in Drug Discovery. Such a method has been developed. It achieves absolute purity and accurate MEC determination by a single automated HPLC analysis that uses less than 5μg of material. Specifically, analytical HPLC separation with online UV detection was used to resolve impurities and measure absorbance from only the compound of interest. Simultaneously, an online chemiluminescence nitrogen detector (CLND) was used to determine the concentration of the analyte. The MECs were then calculated from the absorbance and concentration results. The accuracy of the method was demonstrated using caffeine and a commercial cytotoxic drug, DM1. This approach is particularly suited to analyzing mixtures or samples with low purities. Excellent reproducibility was demonstrated by analyzing a proprietary drug with linker synthesized from different batches with very different levels of purity. In addition, the MECs of drug with linker, along with ADC peak areas measured from size exclusion chromatography (SEC), were used to calculate DARs for 21 in-house ADCs. The DAR results were consistent with those obtained by MS analysis. PMID:27286648

  2. A pathologist-in-the-loop IHC antibody test selection using the entropy-based probabilistic method

    PubMed Central

    Shin, Dmitriy; Arthur, Gerald; Caldwell, Charles; Popescu, Mihail; Petruc, Marius; Diaz-Arias, Alberto; Shyu, Chi-Ren

    2012-01-01

    Background: Immunohistochemistry (IHC) is an important tool to identify and quantify expression of certain proteins (antigens) to gain insights into the molecular processes in a diseased tissue. However, it is a challenge for pathologists to remember the discriminative characteristics of the growing number of such antigens across multiple diseases. The complexity of their expression patterns, fueled by continuous discoveries in molecular pathology, gives rise to a combinatorial explosion that places an unprecedented burden on a practicing pathologist and therefore increases cost and variability of IHC studies. Materials and Methods: To tackle these issues, we have developed antibody test optimized selection method, a novel informatics tool to help pathologists in improving the IHC antibody selection process. The method uses extensions of Shannon's information entropies and Bayesian probabilities to dynamically build an efficient diagnostic tree. Results: A comparative analysis of our method with the expert and World Health Organization classification guidelines showed that the proposed method brings threefold reduction in number of antibody tests required to reach a diagnostic conclusion. Conclusion: The developed method can significantly streamline the antibody test selection process, decrease associated costs and reduce inter- and intrapathologist variability in IHC decision-making. PMID:22439121

  3. Description of a double centrifugation tube method for concentrating canine platelets

    PubMed Central

    2013-01-01

    Background To evaluate the efficiency of platelet-rich plasma preparations by means of a double centrifugation tube method to obtain platelet-rich canine plasma at a concentration at least 4 times higher than the baseline value and a concentration of white blood cells not exceeding twice the reference range. A complete blood count was carried out for each sample and each concentrate. Whole blood samples were collected from 12 clinically healthy dogs (consenting blood donors). Blood was processed by a double centrifugation tube method to obtain platelet concentrates, which were then analyzed by a flow cytometry haematology system for haemogram. Platelet concentration and white blood cell count were determined in all samples. Results Platelet concentration at least 4 times higher than the baseline value and a white blood cell count not exceeding twice the reference range were obtained respectively in 10 cases out of 12 (83.3%) and 11 cases out of 12 (91.6%). Conclusions This double centrifugation tube method is a relatively simple and inexpensive method for obtaining platelet-rich canine plasma, potentially available for therapeutic use to improve the healing process. PMID:23876182

  4. Streamlined method for parallel identification of single domain antibodies to membrane receptors on whole cells

    PubMed Central

    Rossotti, Martín; Tabares, Sofía; Alfaya, Lucía; Leizagoyen, Carmen; Moron, Gabriel; González-Sapienza, Gualberto

    2015-01-01

    BACKGROUND Owing to their minimal size, high production yield, versatility and robustness, the recombinant variable domain (nanobody) of camelid single chain antibodies are valued affinity reagents for research, diagnostic, and therapeutic applications. While their preparation against purified antigens is straightforward, the generation of nanobodies to difficult targets such as multi-pass or complex membrane cell receptors remains challenging. Here we devised a platform for high throughput identification of nanobodies to cell receptor based on the use of a biotin handle. METHODS Using a biotin-acceptor peptide tag, the in vivo biotinylation of nanobodies in 96 well culture blocks was optimized allowing their parallel analysis by flow cytometry and ELISA, and their direct used for pull-down/MS target identification. RESULTS The potential of this strategy was demonstrated by the selection and characterization of panels of nanobodies to Mac-1 (CD11b/CD18), MHC II and the mouse Ly-5 leukocyte common antigen (CD45) receptors, from a VHH library obtained from a llama immunized with mouse bone marrow derived dendritic cells. By on and off switching of the addition of biotin, the method also allowed the epitope binning of the selected Nbs directly on cells. CONCLUSIONS This strategy streamline the selection of potent nanobodies to complex antigens, and the selected nanobodies constitute ready-to-use biotinylated reagents. GENERAL SIGNIFICANCE This method will accelerate the discovery of nanobodies to cell membrane receptors which comprise the largest group of drug and analytical targets. PMID:25819371

  5. Immunoscintigraphy with indium-111 labeled monoclonal antibodies: The importance of a good display method

    SciTech Connect

    Liehn, J.C.; Hannequin, P.; Nasca, S.; Lebrun, D.; Fernandez-Valoni, A.; Valeyre, J. )

    1989-03-01

    A major drawback of In-111-labeled monoclonal antibodies (MoAb) is the presence of intense liver, renal, and bone marrow nonspecific activity. This makes the display of the images hardly optimal and their visual interpretation difficult. In this study, the intrinsic color scale (which consists of selecting the limits of the color scale as the highest and the lowest pixel value of the image) was compared to a new, simple algorithm for the determination of the limits of the color scale. This algorithm was based on the count density in the iliac crest areas. OC-125 or anti-CEA In-111 MoAb F(ab')2 fragments were used in 32 patients with suspected recurrence of ovarian (19 patients) or colorectal cancer (13 patients). Final diagnosis was assessed by surgery (21 patients), biopsy (five patients), or followup (six patients). A 10-minute abdomino-pelvic anterior view was recorded two days after injection. These views are displayed using the two methods and interpreted by two observers. Using their responses in each quadrant of the pelvis, the authors calculated two ROC curves. The comparison of the ROC curves showed better performances for the new method. For example, for the same specificity (73%), the sensitivity of the new method was significantly better (78% versus 68%). This result confirmed the importance of a good methodology for displaying immunoscintigraphic images.

  6. Efficient method to optimize antibodies using avian leukosis virus display and eukaryotic cells.

    PubMed

    Yu, Changming; Pike, Gennett M; Rinkoski, Tommy A; Correia, Cristina; Kaufmann, Scott H; Federspiel, Mark J

    2015-08-11

    Antibody-based therapeutics have now had success in the clinic. The affinity and specificity of the antibody for the target ligand determines the specificity of therapeutic delivery and off-target side effects. The discovery and optimization of high-affinity antibodies to important therapeutic targets could be significantly improved by the availability of a robust, eukaryotic display technology comparable to phage display that would overcome the protein translation limitations of microorganisms. The use of eukaryotic cells would improve the diversity of the displayed antibodies that can be screened and optimized as well as more seamlessly transition into a large-scale mammalian expression system for clinical production. In this study, we demonstrate that the replication and polypeptide display characteristics of a eukaryotic retrovirus, avian leukosis virus (ALV), offers a robust, eukaryotic version of bacteriophage display. The binding affinity of a model single-chain Fv antibody was optimized by using ALV display, improving affinity >2,000-fold, from micromolar to picomolar levels. We believe ALV display provides an extension to antibody display on microorganisms and offers virus and cell display platforms in a eukaryotic expression system. ALV display should enable an improvement in the diversity of properly processed and functional antibody variants that can be screened and affinity-optimized to improve promising antibody candidates. PMID:26216971

  7. Efficient method to optimize antibodies using avian leukosis virus display and eukaryotic cells

    PubMed Central

    Yu, Changming; Pike, Gennett M.; Rinkoski, Tommy A.; Correia, Cristina; Kaufmann, Scott H.; Federspiel, Mark J.

    2015-01-01

    Antibody-based therapeutics have now had success in the clinic. The affinity and specificity of the antibody for the target ligand determines the specificity of therapeutic delivery and off-target side effects. The discovery and optimization of high-affinity antibodies to important therapeutic targets could be significantly improved by the availability of a robust, eukaryotic display technology comparable to phage display that would overcome the protein translation limitations of microorganisms. The use of eukaryotic cells would improve the diversity of the displayed antibodies that can be screened and optimized as well as more seamlessly transition into a large-scale mammalian expression system for clinical production. In this study, we demonstrate that the replication and polypeptide display characteristics of a eukaryotic retrovirus, avian leukosis virus (ALV), offers a robust, eukaryotic version of bacteriophage display. The binding affinity of a model single-chain Fv antibody was optimized by using ALV display, improving affinity >2,000-fold, from micromolar to picomolar levels. We believe ALV display provides an extension to antibody display on microorganisms and offers virus and cell display platforms in a eukaryotic expression system. ALV display should enable an improvement in the diversity of properly processed and functional antibody variants that can be screened and affinity-optimized to improve promising antibody candidates. PMID:26216971

  8. Method and cell lines for the production of monoclonal antibodies to human glycophorin A

    DOEpatents

    Bigbee, W.L.; Fong, S.S.N.; Jensen, R.H.; Vanderlaan, M.

    Cloned mouse hybridoma cell lines have been established which continuously produce antibodies that differentiate between the M and N forms of human glycophorin A. These antibodies have potential application as human blood group reagents, as markers for terminally differentiated erythroid cells and as immunofluorescent labels of somatically variant human erythrocytes.

  9. Novel Microdilution Method to Assess Double and Triple Antibiotic Combination Therapy In Vitro

    PubMed Central

    El-Azizi, Mohamed

    2016-01-01

    An in vitro microdilution method was developed to assess double and triple combinations of antibiotics. Five antibiotics including ciprofloxacin, amikacin, ceftazidime, piperacillin, and imipenem were tested against 10 clinical isolates of Pseudomonas aeruginosa. Each isolate was tested against ten double and nine triple combinations of the antibiotics. A 96-well plate was used to test three antibiotics, each one alone and in double and triple combinations against each isolate. The minimum bacteriostatic and bactericidal concentrations in combination were determined with respect to the most potent antibiotic. An Interaction Code (IC) was generated for each combination, where a numerical value was designated based on the 2-fold increase or decrease in the MICs with respect to the most potent antibiotic. The results of the combinations were verified by time-kill assay at constant concentrations of the antibiotics and in a chemostat. Only 13% of the double combinations were synergistic, whereas 5% showed antagonism. Forty-three percent of the triple combinations were synergistic with no antagonism observed, and 100% synergism was observed in combination of ciprofloxacin, amikacin, and ceftazidime. The presented protocol is simple and fast and can help the clinicians in the early selection of the effective antibiotic therapy for treatment of severe infections. PMID:27195009

  10. Temperature measurement of wood flame based on the double line method of atomic emission spectra

    NASA Astrophysics Data System (ADS)

    Hao, Xiaojian; Liu, Zhenhua; Sang, Tao

    2016-01-01

    Aimed at the testing requirement of the transient high temperature in explosion field and the bore of barrel weapon, the temperature measurement system of double line of atomic emission spectrum was designed, the method of flame spectrum testing system were used for experimental analysis. The experimental study of wood burning spectra was done with flame spectrum testing system. The measured spectra contained atomic emission spectra of the elements K, Na, and the excitation ease of two kinds atomic emission spectra was analyzed. The temperature was calculated with two spectral lines of K I 766.5nm and 769.9nm. The results show that, compared with Na, the excitation temperature of K atomic emission spectra is lower. By double line method, the temperature of wood burning is 1040K, and error is 3.7%.

  11. Research on text encryption and hiding method with double-random phase-encoding

    NASA Astrophysics Data System (ADS)

    Xu, Hongsheng; Sang, Nong

    2013-10-01

    By using optical image processing techniques, a novel text encryption and hiding method applied by double-random phase-encoding technique is proposed in the paper. The first step is that the secret message is transformed into a 2- dimension array. The higher bits of the elements in the array are used to fill with the bit stream of the secret text, while the lower bits are stored specific values. Then, the transformed array is encoded by double random phase encoding technique. Last, the encoded array is embedded on a public host image to obtain the image embedded with hidden text. The performance of the proposed technique is tested via analytical modeling and test data stream. Experimental results show that the secret text can be recovered either accurately or almost accurately, while maintaining the quality of the host image embedded with hidden data by properly selecting the method of transforming the secret text into an array and the superimposition coefficient.

  12. Chosen-plaintext attack on double-random-phase-encoding-based image hiding method

    NASA Astrophysics Data System (ADS)

    Xu, Hongsheng; Li, Guirong; Zhu, Xianchen

    2015-12-01

    By using optical image processing techniques, a novel text encryption and hiding method applied by double-random phase-encoding technique is proposed in the paper. The first step is that the secret message is transformed into a 2- dimension array. The higher bits of the elements in the array are used to fill with the bit stream of the secret text, while the lower bits are stored specific values. Then, the transformed array is encoded by double random phase encoding technique. Last, the encoded array is embedded on a public host image to obtain the image embedded with hidden text. The performance of the proposed technique is tested via analytical modeling and test data stream. Experimental results show that the secret text can be recovered either accurately or almost accurately, while maintaining the quality of the host image embedded with hidden data by properly selecting the method of transforming the secret text into an array and the superimposition coefficient.

  13. Key management of the double random-phase-encoding method using public-key encryption

    NASA Astrophysics Data System (ADS)

    Saini, Nirmala; Sinha, Aloka

    2010-03-01

    Public-key encryption has been used to encode the key of the encryption process. In the proposed technique, an input image has been encrypted by using the double random-phase-encoding method using extended fractional Fourier transform. The key of the encryption process have been encoded by using the Rivest-Shamir-Adelman (RSA) public-key encryption algorithm. The encoded key has then been transmitted to the receiver side along with the encrypted image. In the decryption process, first the encoded key has been decrypted using the secret key and then the encrypted image has been decrypted by using the retrieved key parameters. The proposed technique has advantage over double random-phase-encoding method because the problem associated with the transmission of the key has been eliminated by using public-key encryption. Computer simulation has been carried out to validate the proposed technique.

  14. Double tracer autoradiographic method for sequential evaluation of regional cerebral perfusion

    SciTech Connect

    Matsuda, H.; Tsuji, S.; Oba, H.; Kinuya, K.; Terada, H.; Sumiya, H.; Shiba, K.; Mori, H.; Hisada, K.; Maeda, T. )

    1989-01-01

    A new double tracer autoradiographic method for the sequential evaluation of altered regional cerebral perfusion in the same animal is presented. This method is based on the sequential injection of two tracers, {sup 99m}Tc-hexamethylpropyleneamine oxime and N-isopropyl-({sup 125}I)p-iodoamphetamine. This method is validated in the assessment of brovincamine effects on regional cerebral perfusion in an experimental model of chronic brain ischemia in the rat. The drug enhanced perfusion recovery in low-flow areas, selectively in surrounding areas of infarction. The results suggest that this technique is of potential use in the study of neuropharmacological effects applied during the experiment.

  15. A Double-difference Earthquake location algorithm: Method and application to the Northern Hayward Fault, California

    USGS Publications Warehouse

    Waldhauser, F.; Ellsworth, W.L.

    2000-01-01

    We have developed an efficient method to determine high-resolution hypocenter locations over large distances. The location method incorporates ordinary absolute travel-time measurements and/or cross-correlation P-and S-wave differential travel-time measurements. Residuals between observed and theoretical travel-time differences (or double-differences) are minimized for pairs of earthquakes at each station while linking together all observed event-station pairs. A least-squares solution is found by iteratively adjusting the vector difference between hypocentral pairs. The double-difference algorithm minimizes errors due to unmodeled velocity structure without the use of station corrections. Because catalog and cross-correlation data are combined into one system of equations, interevent distances within multiplets are determined to the accuracy of the cross-correlation data, while the relative locations between multiplets and uncorrelated events are simultaneously determined to the accuracy of the absolute travel-time data. Statistical resampling methods are used to estimate data accuracy and location errors. Uncertainties in double-difference locations are improved by more than an order of magnitude compared to catalog locations. The algorithm is tested, and its performance is demonstrated on two clusters of earthquakes located on the northern Hayward fault, California. There it colapses the diffuse catalog locations into sharp images of seismicity and reveals horizontal lineations of hypocenter that define the narrow regions on the fault where stress is released by brittle failure.

  16. Cell Proliferation Method: Click Chemistry Based on BrdU Coupling for Multiplex Antibody Staining.

    PubMed

    Cappella, Paolo; Gasparri, Fabio; Pulici, Maurizio; Moll, Jürgen

    2015-01-01

    Determination of incorporation of the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) into DNA is a widely used method to analyze the cell cycle. However, DNA denaturation is required for BrdU detection with the consequence that most protein epitopes are destroyed and their immunocytochemical detection for multiplex analysis is not possible. A novel assay is presented for identifying cells in active S-phase that does not require the DNA denaturation step but nevertheless detects BrdU. For this purpose, cells were pulsed for a short time by 5-ethynyl-2'-deoxyuridine (EdU) which is incorporated into DNA. The nucleotide-exposed ethynyl residue was then derivatized by a copper-catalyzed cycloaddition reaction ("click chemistry" coupling) using a BrdU azide probe. The resulting DNA-bound bromouracil moieties were then detected by commercial anti-BrdU monoclonal antibodies without the need for a denaturation step. This method has been tested using several cell lines and is more sensitive than traditional BrdU and allows multicolor and multiplex analysis in flow cytometry (FCM) and image-based cytometry. PMID:25827484

  17. Detection of Infertility-related Neutralizing Antibodies with a Cell-free Microfluidic Method

    NASA Astrophysics Data System (ADS)

    Eyer, Klaus; Root, Katharina; Verboket, Pascal E.; Dittrich, Petra S.

    2015-11-01

    The unwanted emergence of neutralizing antibodies (nAbs) against an endogenous or a therapeutic protein can result in deficiency diseases or therapy failure. Here, we developed a cell-free microfluidic method for the sensitive detection and quantification of nAbs in human serum that are associated with infertility. We used cell-derived vesicles containing the luteinizing hormone (LH)/choriogonadotropin receptor (LHHCGR) to detect nAbs against LH. The method exploits the entire cellular signal amplification mechanism, and facilitates the detection of as little as 0.44 nM of LH-nAb (Kd 1.5 nM) in human serum matrix within only 15 minutes. In addition, dose-response curves can be generated in less than 2 hours to evaluate the nAB concentration and dissociation constant. The developed system is devoid of problems associated with cell-based assays and we believe that this simple effect-directed analysis can be used in clinical environments, and is adaptable to other hormones or cytokines and their respective nAbs.

  18. Cell proliferation method: click chemistry based on BrdU coupling for multiplex antibody staining.

    PubMed

    Cappella, Paolo; Gasparri, Fabio; Pulici, Maurizio; Moll, Jürgen

    2008-07-01

    Determination of incorporation of the thymidine analog 5-bromo-2'-deoxyuridine (BrdU) into DNA is a widely used method to analyze the cell cycle (see UNIT 7.7). However, DNA denaturation is required for BrdU detection with the consequence that most protein epitopes are destroyed and their immunocytochemical detection for multiplex analysis is not possible. A novel assay is presented for identifying cells in active S-phase that does not require the DNA denaturation step but nevertheless detects BrdU. For this purpose, cells were pulsed for a short time by an alkenyl deoxyuridine (5-ethynyl-2'-deoxyuridine, EdU), which is incorporated into DNA. The nucleotide exposed ethynyl residue was then derivatized by a copper-catalyzed cycloaddition reaction ("click chemistry" coupling) using a BrdU azide probe. The resulting DNA-bound bromouracil moieties were then detected by commercial anti-BrdU monoclonal antibodies without the need for a denaturation step. This method has been tested using several cell lines and is preferred over traditional BrdU detection since it is more sensitive and allows multicolor and multiplex analysis in FCM and imaging. PMID:18770651

  19. Detection of Infertility-related Neutralizing Antibodies with a Cell-free Microfluidic Method

    PubMed Central

    Eyer, Klaus; Root, Katharina; Verboket, Pascal E.; Dittrich, Petra S.

    2015-01-01

    The unwanted emergence of neutralizing antibodies (nAbs) against an endogenous or a therapeutic protein can result in deficiency diseases or therapy failure. Here, we developed a cell-free microfluidic method for the sensitive detection and quantification of nAbs in human serum that are associated with infertility. We used cell-derived vesicles containing the luteinizing hormone (LH)/choriogonadotropin receptor (LHHCGR) to detect nAbs against LH. The method exploits the entire cellular signal amplification mechanism, and facilitates the detection of as little as 0.44 nM of LH-nAb (Kd 1.5 nM) in human serum matrix within only 15 minutes. In addition, dose-response curves can be generated in less than 2 hours to evaluate the nAB concentration and dissociation constant. The developed system is devoid of problems associated with cell-based assays and we believe that this simple effect-directed analysis can be used in clinical environments, and is adaptable to other hormones or cytokines and their respective nAbs. PMID:26585778

  20. High resolution image reconstruction method for a double-plane PET system with changeable spacing

    NASA Astrophysics Data System (ADS)

    Gu, Xiao-Yue; Zhou, Wei; Li, Lin; Wei, Long; Yin, Peng-Fei; Shang, Lei-Min; Yun, Ming-Kai; Lu, Zhen-Rui; Huang, Xian-Chao

    2016-05-01

    Breast-dedicated positron emission tomography (PET) imaging techniques have been developed in recent years. Their capacities to detect millimeter-sized breast tumors have been the subject of many studies. Some of them have been confirmed with good results in clinical applications. With regard to biopsy application, a double-plane detector arrangement is practicable, as it offers the convenience of breast immobilization. However, the serious blurring effect of the double-plane PET, with changeable spacing for different breast sizes, should be studied. We investigated a high resolution reconstruction method applicable for a double-plane PET. The distance between the detector planes is changeable. Geometric and blurring components were calculated in real-time for different detector distances, and accurate geometric sensitivity was obtained with a new tube area model. Resolution recovery was achieved by estimating blurring effects derived from simulated single gamma response information. The results showed that the new geometric modeling gave a more finite and smooth sensitivity weight in the double-plane PET. The blurring component yielded contrast recovery levels that could not be reached without blurring modeling, and improved visual recovery of the smallest spheres and better delineation of the structures in the reconstructed images were achieved with the blurring component. Statistical noise had lower variance at the voxel level with blurring modeling at matched resolution, compared to without blurring modeling. In distance-changeable double-plane PET, finite resolution modeling during reconstruction achieved resolution recovery, without noise amplification. Supported by Knowledge Innovation Project of The Chinese Academy of Sciences (KJCX2-EW-N06)

  1. A Simple Method for the Immunocytochemical Detection of Proteins Inside Nuclear Structures That Are Inaccessible to Specific Antibodies

    PubMed Central

    Svistunova, Darya M.; Musinova, Yana R.; Polyakov, Vladimir Yu.

    2012-01-01

    It has been demonstrated elsewhere that a high concentration of an antigen within the nucleolus may prevent its proper recognition by specific antibodies. In this study, the authors found that a short proteinase treatment allowed for the detection of antigens in the nucleoli. The described approach is compatible with the simultaneous observation of proteins fused to fluorescent tags and with preembedding electron microscopy. It appears that the described method can be useful in situations when the proper recognition of antigens by specific antibodies is disturbed by a high density of cellular structures or a high concentration of antigens inside these structures. PMID:22114257

  2. A novel method of preparing the monoform structure of catalytic antibody light chain.

    PubMed

    Hifumi, Emi; Matsumoto, Shingo; Nakashima, Hiroki; Itonaga, Shogo; Arakawa, Mitsue; Katayama, Yoshiki; Kato, Ryuichi; Uda, Taizo

    2016-02-01

    Along with the development of antibody drugs and catalytic antibodies, the structural diversity (heterogeneity) of antibodies has been given attention. For >20 yr, detailed studies on the subject have not been conducted, because the phenomenon presents many difficult and complex problems. Structural diversity provides some (or many) isoforms of an antibody distinguished by different charges, different molecular sizes, and modifications of amino acid residues. For practical use, the antibody and the subunits must have a defined structure. In recent work, we have found that the copper (Cu) ion plays a substantial role in solving the diversity problem. In the current study, we used several catalytic antibody light chains to examine the effect of the Cu ion. In all cases, the different electrical charges of the molecule converged to a single charge, giving 1 peak in cation-exchange chromatography, as well as a single spot in 2-dimensional gel electrophoresis. The Cu-binding site was investigated by using mutagenesis, ultraviolet-visible spectroscopy, atomic force microscope analysis, and molecular modeling, which suggested that histidine and cysteine residues close to the C-terminus are involved with the binding site. The constant region domain of the antibody light chain played an important role in the heterogeneity of the light chain. Our findings may be a significant tool for preparing a single defined, not multiple, isoform structure. PMID:26527062

  3. Targeting Aberrant DNA double strand break repair in triple negative breast cancer with alpha particle emitter radiolabeled anti-EGFR antibody

    PubMed Central

    Song, Hong; Hedayati, Mohammad; Hobbs, Robert F.; Shao, Chunbo; Bruchertseifer, Frank; Morgenstern, Alfred; DeWeese, Theodore L.; Sgouros, George

    2013-01-01

    The higher potential efficacy of alpha-particle radiopharmaceutical therapy lies in the 3 to 8-fold greater biological effectiveness (RBE) of alpha particles relative to photon or beta-particle radiation. This greater RBE, however, also applies to normal tissue, thereby reducing the potential advantage of high RBE. Since alpha particles typically cause DNA double strand breaks (DSBs), targeting tumors that are defective in DSB repair effectively increases the RBE, yielding a secondary, RBE-based differentiation between tumor and normal tissue that is complementary to conventional, receptor-mediated tumor targeting. In some triple negative breast cancers (TNBC, ER−/PR−/HER-2−), germline mutation in BRCA-1, a key gene in homologous recombination (HR) DSB repair, predisposes patients to early onset of breast cancer. These patients have few treatment options once the cancer has metastasized. In this study, we investigated the efficacy of alpha particle emitter, 213Bi labeled anti-EGFR antibody, Cetuximab, in BRCA-1 defective TNBC. 213Bi-Cetuximab was found to be significantly more effective in the BRCA-1 mutated TNBC cell line HCC1937 than BRCA-1 competent TNBC cell MDA-MB-231. siRNA knockdown of BRCA-1 or DNA-PKcs, a key gene in non-homologous end joining (NHEJ) DSB repair pathway, also sensitized TNBC cells to 213Bi-Cetuximab. Furthermore, the small molecule inhibitor of DNA-PKcs, NU7441, sensitized BRCA-1 competent TNBC cells to alpha particle radiation. Immunofluorescent staining of γH2AX foci and comet assay confirmed that enhanced RBE is caused by impaired DSB repair. These data offer a novel strategy for enhancing conventional receptor-mediated targeting with an additional, potentially synergistic radiobiological targeting that could be applied to TNBC. PMID:23873849

  4. Simple NMR methods for evaluating higher order structures of monoclonal antibody therapeutics with quinary structure.

    PubMed

    Chen, Kang; Long, Dianna S; Lute, Scott C; Levy, Michaella J; Brorson, Kurt A; Keire, David A

    2016-09-01

    Monoclonal antibody (mAb) drugs constitute the largest class of protein therapeutics currently on the market. Correctly folded protein higher order structure (HOS), including quinary structure, is crucial for mAb drug quality. The quinary structure is defined as the association of quaternary structures (e.g., oligomerized mAb). Here, several commonly available analytical methods, i.e., size-exclusion-chromatography (SEC) FPLC, multi-angle light scattering (MALS), circular dichroism (CD), NMR and multivariate analysis, were combined and modified to yield a complete profile of HOS and comparable metrics. Rituximab and infliximab were chosen for method evaluation because both IgG1 molecules are known to be homologous in sequence, superimposable in Fab crystal structure and identical in Fc structure. However, herein the two are identified to be significantly different in quinary structure in addition to minor secondary structure differences. All data collectively showed rituximab was mostly monomeric while infliximab was in mono-oligomer equilibrium driven by its Fab fragment. The quinary structure differences were qualitatively inferred from the less used but more reproducible dilution-injection-SEC-FPLC curve method. Quantitative principal component analysis (PCA) was performed on NMR spectra of either the intact or the in-situ enzymatic-digested mAb samples. The cleavage reactions happened directly in NMR tubes without further separation, which greatly enhanced NMR spectra quality and resulted in larger inter- and intra-lot variations based on PCA. The new in-situ enzymatic digestion method holds potential in identifying structural differences on larger therapeutic molecules using NMR. PMID:27344629

  5. Single-Mach and double-Mach reflection - Its representation in Ernst Mach's historical soot method

    NASA Astrophysics Data System (ADS)

    Krehl, P.

    In 1875 Ernst Mach discovered the effect of irregular interaction of shock waves, the so-called single Mach reflection (SMR), which for symmetric geometry is characterized by two triple points. He recorded their two trajectories on a soot-covered glass plate. Appearing as two mirror-symmetric V-branches, they form the well-known Mach soot funnel. Combining this soot method with the schlieren technique facilitates the interpretation of soot-recorded interaction phenomena as well as allows to resolve the soot removal mechanism in time. Increasing the dynamic recording range of the soot layer in terms of reflected shock pressures even renders visualization of double-Mach reflection (DMR) which, in the case of symmetric shock interaction, is characterized by a second concentric, external 'double-Mach funnel'. At transition of DMR to SMR it merges into the ordinary 'single-Mach funnel'.

  6. Earthquake hypocenter relocation using double difference method in East Java and surrounding areas

    SciTech Connect

    C, Aprilia Puspita; Nugraha, Andri Dian; Puspito, Nanang T

    2015-04-24

    Determination of precise hypocenter location is very important in order to provide information about subsurface fault plane and for seismic hazard analysis. In this study, we have relocated hypocenter earthquakes in Eastern part of Java and surrounding areas from local earthquake data catalog compiled by Meteorological, Climatological, and Geophysical Agency of Indonesia (MCGA) in time period 2009-2012 by using the double-difference method. The results show that after relocation processes, there are significantly changes in position and orientation of earthquake hypocenter which is correlated with the geological setting in this region. We observed indication of double seismic zone at depths of 70-120 km within the subducting slab in south of eastern part of Java region. Our results will provide useful information for advance seismological studies and seismic hazard analysis in this study.

  7. Earthquake hypocenter relocation using double difference method in East Java and surrounding areas

    NASA Astrophysics Data System (ADS)

    C, Aprilia Puspita; Nugraha, Andri Dian; Puspito, Nanang T.

    2015-04-01

    Determination of precise hypocenter location is very important in order to provide information about subsurface fault plane and for seismic hazard analysis. In this study, we have relocated hypocenter earthquakes in Eastern part of Java and surrounding areas from local earthquake data catalog compiled by Meteorological, Climatological, and Geophysical Agency of Indonesia (MCGA) in time period 2009-2012 by using the double-difference method. The results show that after relocation processes, there are significantly changes in position and orientation of earthquake hypocenter which is correlated with the geological setting in this region. We observed indication of double seismic zone at depths of 70-120 km within the subducting slab in south of eastern part of Java region. Our results will provide useful information for advance seismological studies and seismic hazard analysis in this study.

  8. Improved Methods to Detect Low Levels of HIV Using Antibody-Based Technologies.

    PubMed

    Eugenin, Eliseo A; Berman, Joan W

    2016-01-01

    Persistence of latent virus represents a major barrier to eradicating HIV even in the current antiretroviral therapy era. A critical limitation to eliminating these viral reservoirs is the lack of reliable methods to detect, quantify, and characterize cells harboring low levels of virus. However, recent work of several laboratories indicates that PCR and viral amplification based technologies underestimate or overestimate the size of the reservoirs. Thus, new technologies and methodologies to detect, quantify, and characterize these viral reservoirs are necessary to monitor and eradicate HIV. Recent developments in imaging technologies have enabled the development or improvement of detection protocols and have facilitated the identification and quantification of several markers with exquisite resolution. In the context of HIV, we developed new protocols for the detection of low amounts of viral proteins. In this chapter, we describe several antibody-based technologies for signal amplification to improve and detect low amounts of HIV proteins in cells, tissues, and other biological samples. The improvement in these techniques is essential to detect viral reservoirs and to design strategies to eliminate them. PMID:26714718

  9. Low-noise multiple watermarks technology based on complex double random phase encoding method

    NASA Astrophysics Data System (ADS)

    Zheng, Jihong; Lu, Rongwen; Sun, Liujie; Zhuang, Songlin

    2010-11-01

    Based on double random phase encoding method (DRPE), watermarking technology may provide a stable and robust method to protect the copyright of the printing. However, due to its linear character, DRPE exist the serious safety risk when it is attacked. In this paper, a complex coding method, which means adding the chaotic encryption based on logistic mapping before the DRPE coding, is provided and simulated. The results testify the complex method will provide better security protection for the watermarking. Furthermore, a low-noise multiple watermarking is studied, which means embedding multiple watermarks into one host printing and decrypt them with corresponding phase keys individually. The Digital simulation and mathematic analysis show that with the same total embedding weight factor, multiply watermarking will improve signal noise ratio (SNR) of the output printing image significantly. The complex multiply watermark method may provide a robust, stability, reliability copyright protection with higher quality printing image.

  10. Hypersingular meshless method using double-layer potentials for three-dimensional exterior acoustic problems.

    PubMed

    Young, D L; Chen, K H; Liu, T Y; Wu, C S

    2016-01-01

    Three-dimensional exterior acoustic problems with irregular domains are solved using a hypersingular meshless method. In particular, the method of fundamental solutions (MFS) is used to formulate and analyze such acoustic problems. It is well known that source points for MFS cannot be located on the real boundary due to the singularity of the kernel functions. Thus, the diagonal terms of the influence matrices are unobtainable when source points are located on the boundary. An efficient approach is proposed to overcome such difficulties, when the MFS is used for three-dimensional exterior acoustic problems. This work is an extension of previous research on two-dimensional problems. The solution of the problem is expressed in terms of a double-layer potential representation on the physical boundary. Three examples are presented in which the proposed method is compared to the MFS and boundary element method. Good numerical performance is demonstrated by the proposed hypersingular meshless method. PMID:26827046

  11. A time-resolved Langmuir double-probe method for the investigation of pulsed magnetron discharges

    SciTech Connect

    Welzel, Th.; Dunger, Th.; Kupfer, H.; Richter, F.

    2004-12-15

    Langmuir probes are important means for the characterization of plasma discharges. For measurements in plasmas used for the deposition of thin films, the Langmuir double probe is especially suited. With the increasing popularity of pulsed deposition discharges, there is also an increasing need for time-resolved characterization methods. For Langmuir probes, several single-probe approaches to time-resolved measurements are reported but very few for the double probe. We present a time-resolved Langmuir double-probe technique, which is applied to a pulsed magnetron discharge at several 100 kHz used for MgO deposition. The investigations show that a proper treatment of the current measurement is necessary to obtain reliable results. In doing so, a characteristic time dependence of the charge-carrier density during the ''pulse on'' time containing maximum values of almost 2{center_dot}10{sup 11} cm{sup -3} was found. This characteristic time dependence varies with the pulse frequency and the duty cycle. A similar time dependence of the electron temperature is only observed when the probe is placed near the magnesium target.

  12. Second-order perturbation corrections to singles and doubles coupled-cluster methods: General theory and application to the valence optimized doubles model

    NASA Astrophysics Data System (ADS)

    Gwaltney, Steven R.; Sherrill, C. David; Head-Gordon, Martin; Krylov, Anna I.

    2000-09-01

    We present a general perturbative method for correcting a singles and doubles coupled-cluster energy. The coupled-cluster wave function is used to define a similarity-transformed Hamiltonian, which is partitioned into a zeroth-order part that the reference problem solves exactly plus a first-order perturbation. Standard perturbation theory through second-order provides the leading correction. Applied to the valence optimized doubles (VOD) approximation to the full-valence complete active space self-consistent field method, the second-order correction, which we call (2), captures dynamical correlation effects through external single, double, and semi-internal triple and quadruple substitutions. A factorization approximation reduces the cost of the quadruple substitutions to only sixth order in the size of the molecule. A series of numerical tests are presented showing that VOD(2) is stable and well-behaved provided that the VOD reference is also stable. The second-order correction is also general to standard unwindowed coupled-cluster energies such as the coupled-cluster singles and doubles (CCSD) method itself, and the equations presented here fully define the corresponding CCSD(2) energy.

  13. 349 Detection of Anti-nucclear Antibodies (ana) Used for Diagnostic Approach of Systemic Autoimmune Diseases. Correlation with Double Stranded DNA (DSDNA) and Extractable Nuclear Antigen (ENA) Antibodies

    PubMed Central

    Anastasiou, Ekarerini; Vakaloudi, Anastasia; Papadopoulos, Georgios; Mavridou, Styliani; Koteli, Asimoula

    2012-01-01

    Background To determine the correlation between the titer of ANA and anti-dsDNA and anti-ENA antibodies and the contribution of ANA detection to the diagnosis of connective tissue diseases (CTD). Methods Our samples consisted of 516 specimens, from Rheumatology Department, collected during January 2010 – July 2010. The detection of ANA was performed using indirect immunofluorescence (IFA) and the detection of anti-dsDNA and anti-ENA using ELISA. Results Of the 364 (70.54%) samples with negative ANA 4 (1%) had positive anti-ENA and 2 (0.5%) had positive anti-dsDNA while positive anti-ENA and anti-dsDNA were detected in the 44.73% (n = 68) and 21% (n = 32) of the specimens with positive ANA respectively. The probability of detecting positive anti-ENA and anti-dsDNA rises proportionately to the titer of ANA. Specifically, the correlation between the probability of detecting positive anti-ENA and the titer of ANA is 0.577 (P < 0.001) while the correlation between the probability of detecting positive anti-dsDNA and the titer of ANA is 0.18 (P = 0.003). Probability calculations on the basis of the ANA titer showed that samples with low titer ANAs (1:160 or less) had low probabilities for positive anti-ENA. The receiver operating (ROC) curves of the ANA titer for anti-ENA had a larger under the curve area compared to the ROC curve for anti-dsDNA, indicating that ANA titer is better for predicting anti-ENA than anti-dsDNA. The sensitivity of positive ANA in the prediction of the anti-ENA and anti-dsDNA was 94.40% and 94.10%, the specificity was 81% and 75.10%, the positive prognostic value was 44.70% and 21.10% and negative prognostic value was 98.90 and 99.50%. Conclusions The detection of ANA using indirect IFA has high sensitivity in predicting the presence of specialized antibodies and may be used as a screening method for the diagnosis of CTD. It is cost and time effective too. Our study also shows that the ANA titer is useful in predicting anti-ENA. Samples with

  14. A new virus discovered by immunocapture of double-stranded RNA, a rapid method for virus enrichment in metagenomic studies.

    PubMed

    Blouin, Arnaud G; Ross, Howard A; Hobson-Peters, Jody; O'Brien, Caitlin A; Warren, Ben; MacDiarmid, Robin

    2016-09-01

    Next-generation sequencing technologies enable the rapid identification of viral infection of diseased organisms. However, despite a consistent decrease in sequencing costs, it is difficult to justify their use in large-scale surveys without a virus sequence enrichment technique. As the majority of plant viruses have an RNA genome, a common approach is to extract the double-stranded RNA (dsRNA) replicative form, to enrich the replicating virus genetic material over the host background. The traditional dsRNA extraction is time-consuming and labour-intensive. We present an alternative method to enrich dsRNA from plant extracts using anti-dsRNA monoclonal antibodies in a pull-down assay. The extracted dsRNA can be amplified by reverse transcriptase-polymerase chain reaction and sequenced by next-generation sequencing. In our study, we have selected three distinct plant hosts: Māori potato (Solanum tuberosum), rengarenga (Arthropodium cirratum) and broadleaved dock (Rumex obtusifolius) representing a cultivated crop, a New Zealand-native ornamental plant and a weed, respectively. Of the sequence data obtained, 31-74% of the reads were of viral origin, and we identified five viruses including Potato virus Y and Potato virus S in potato; Turnip mosaic virus in rengarenga (a new host record); and in the dock sample Cherry leaf roll virus and a novel virus belonging to the genus Macluravirus. We believe that this new assay represents a significant opportunity to upscale virus ecology studies from environmental, primary industry and/or medical samples. PMID:26990372

  15. Sensitivity of a radioimmunoassay method for detection of certain viral antibodies in sera and cerebrospinal fluids.

    PubMed Central

    Forghani, B; Schmidt, N J; Lennette, E H

    1976-01-01

    An indirect solid-phase radioimmunoassay (RIA) was applied to titration of serum and cerebrospinal fluid (CSF) antibodies against a variety of viruses including rubella, mumps, measles, herpes simplex, varicella-zoster, and vaccinia. The test used fixed, virus-infected cells as a source of antigen, and conditions for optimal production of viral antigen were determined for each virus-host cell system. In acute, uncomplicated viral infections, sera taken 2 to 5 days after onset generally had low homotypic RIA titers ranging from less than 1:100 to 1:500, whereas convalescent-phase titers ranged from 1:128,000 to 1:512,000. Rubella and measles antibody titers as high as 1:256,000 were demonstrated by RIA in CSF from patients with chronic panencephalitis, whereas homologous antibody titers of 1:4,000 were detected in CSF from acute mumps, herpes simplex, and varicella-zoster virus infections with central nervous system involvement. Some heterotypic antibody was demonstrable by RIA in CSF, but, with the exception of herpes simplex antibody in a mumps virus infection, titers were markedly lower than those to the infecting virus type. RIA generally demonstrated titers at least 1,000 times higher than those obtained by conventional assays such as complement fixation, hemagglutination inhibition, neutralization, and immunofluorescent staining. PMID:187618

  16. Estimates of Mumps Seroprevalence May Be Influenced by Antibody Specificity and Serologic Method

    PubMed Central

    McGrew, Marcia; Williams, Nobia J.; Sowers, Sun B.; Bellini, William J.; Hickman, Carole J.

    2014-01-01

    Neutralizing antibodies are assumed to be essential for protection against mumps virus infection, but their measurement is labor- and time-intensive. For this reason, enzyme-linked immunosorbent assays (ELISAs) are typically used to measure mumps-specific IgG levels. However, since there is poor correlation between mumps neutralization titers and ELISAs that measure the presence of mumps-specific IgG levels, ELISAs that better correlate with neutralization are needed. To address this issue, we measured mumps antibody levels by plaque reduction neutralization, by a commercial ELISA (whole-virus antigen), and by ELISAs specific for the mumps nucleoprotein and hemagglutinin. The results indicate that differences in the antibody response to the individual mumps proteins could partially explain the lack of correlation among various serologic tests. Furthermore, the data indicate that some seropositive individuals have low levels of neutralizing antibody. If neutralizing antibody is important for protection, this suggests that previous estimates of immunity based on whole-virus ELISAs may be overstated. PMID:24371258

  17. Methods for quantitative detection of antibody-induced complement activation on red blood cells.

    PubMed

    Meulenbroek, Elisabeth M; Wouters, Diana; Zeerleder, Sacha

    2014-01-01

    Antibodies against red blood cells (RBCs) can lead to complement activation resulting in an accelerated clearance via complement receptors in the liver (extravascular hemolysis) or leading to intravascular lysis of RBCs. Alloantibodies (e.g. ABO) or autoantibodies to RBC antigens (as seen in autoimmune hemolytic anemia, AIHA) leading to complement activation are potentially harmful and can be - especially when leading to intravascular lysis - fatal(1). Currently, complement activation due to (auto)-antibodies on RBCs is assessed in vitro by using the Coombs test reflecting complement deposition on RBC or by a nonquantitative hemolytic assay reflecting RBC lysis(1-4). However, to assess the efficacy of complement inhibitors, it is mandatory to have quantitative techniques. Here we describe two such techniques. First, an assay to detect C3 and C4 deposition on red blood cells that is induced by antibodies in patient serum is presented. For this, FACS analysis is used with fluorescently labeled anti-C3 or anti-C4 antibodies. Next, a quantitative hemolytic assay is described. In this assay, complement-mediated hemolysis induced by patient serum is measured making use of spectrophotometric detection of the released hemoglobin. Both of these assays are very reproducible and quantitative, facilitating studies of antibody-induced complement activation. PMID:24514151

  18. Investigation of lasing in lead vapor by the double pulse method

    SciTech Connect

    Kazakov, V.V.; Markova, S.V.; Molchanova, L.V.; Petrash, G.G.

    1983-05-01

    The double pulse method was to study the 722.9 nm line emitted by a lead vapor laser. The parameters of the second excitation pulse were measured simultaneously with the lasing characteristics. It was found that the main factor determining a decrease in the laser output power in the second pulse was the residual population of the lower active level. Variations in the excitation pulse had a relatively weak influence on the form of this dependence and were essentially manifested when there was a relatively long delay between the excitation pulses.

  19. Double-filtering method based on two acousto-optic tunable filters for hyperspectral imaging application.

    PubMed

    Wang, Pengchong; Zhang, Zhonghua

    2016-05-01

    A hyperspectral imaging system was demonstrated based on two acousto-optic tunable filters (AOTFs). Efficient regulation of the incoherent beam was executed by means of the wide-angular regime of Bragg diffraction in the birefringent materials. A double-filtering process was achieved when these two AOTFs operated with a central wavelength difference. In comparison with the single-filtering method, the spectral bandwidth was greatly compressed, giving an increment of 42.02% in spectral resolution at the wavelength of 651.62 nm. Experimental results and theoretical calculations are basically identical. Furthermore, the sidelobe was found to be suppressed by the double-filtering process with the first order maximum decreased from -9.25 dB to -22.35 dB. The results indicated high spectral resolution and high spectral purity were obtained simultaneously from this method. The basic spectral resolution performance was examined with a didymium glass by this configuration. We present our experimental methods and the detailed results obtained. PMID:27137600

  20. Anti-vascular endothelial cell antibodies (AECA): comparison of two assay methods and clinical applications.

    PubMed

    Meyer, O; Kaiser, P; Haim, T; Edgell, C J; Pasquier, C; de Bandt, M; Bridey, F; Sellak, H; Lansaman, J; Kahn, M F

    1995-12-01

    Vascular endothelial cells may be a target for autoantibodies (AECAs) against membrane antigens that are constitutively expressed, induced or bound to their surface. To test this hypothesis, we used an enzyme-linked immunosorbent assay (ELISA) with two types of human endothelial cells as the substrate, i.e., human umbilical cord vein endothelial cells (HUVECs) or the hybrid cell line EAhy-926 obtained by fusion of HUVECs with the bronchial carcinoma cell line A549. A comparative functional study of these two cell types demonstrated that EAhy-926 cells produced only small amounts of VIII von Willebrand factor and tissular factor, did not contain Weibel Palade bodies visible under the electron microscope, and expressed ICAM-1 and selectin E in levels of no more than 15% of those expressed by human umbilical cord vein endothelial cells both after stimulation by bacterial lipopolysaccharide and under basal conditions. However, the two assay methods yielded similar IgG AECA titers when used on sera from patients with rheumatoid vasculitis or antiphospholipid syndrome. These antibodies did not exhibit cytotoxicity for cord vein or EAhy-926 cells. They were not specific for endothelium, since their activity decreased by a mean of 40% after incubation of sera with the epithelial cell line A549. A cross-sectional study of 565 sera demonstrated that anti-vascular IgG and IgM AECAs reactive with EAhy-926 cells occurred mainly in patients with dermatomyositis (IgG, 58%; IgM, 22%), systemic scleroderma (IgG, 48%; IgM, 18%), primary Sjögren's syndrome (IgG, 44%; IgM, 12%) and secondary and primary systemic vasculitides (IgG, 38%; IgM, 18%) including Wegener's granulomatosis. A longitudinal study in patients with Wegener's granulomatosis showed that AECAS were predictive of disease activity. PMID:8869215

  1. The expression of acidic ribosomal phosphoproteins on the surface membrane of different tissues in autoimmune and normal mice which are the target molecules for anti-double-stranded DNA antibodies.

    PubMed Central

    Sun, K H; Liu, W T; Tang, S J; Tsai, C Y; Hsieh, S C; Wu, T H; Han, S H; Yu, C L

    1996-01-01

    Affinity-purified polyclonal anti-double-stranded DNA (anti-dsDNA) antibodies from patients with systemic lupus erythematosus (SLE) exert a cytostatic effect on cultured rat glomerular mesangial cells (MC). The cognate antigens expressed on the surface of MC have been proved to be acidic ribosomal phosphoproteins (P proteins) in our previous study. The mesangial cytostatic effect of anti-dsDNA antibodies is attributed to the cross-reactivity of the antibodies with membrane-expressed P proteins, but not to the effect of minute amounts of anti-ribosomal P proteins antibodies contained in the anti-dsDNA preparations. Immunofluorescence staining of the native cells demonstrated that anti-dsDNA antibodies bound to the surface of rat mesangial cells, rat brain astrocytes (RBA-1) and mouse fibroblasts (3T3). Anti-dsDNA antibodies also exert potent cytostatic effects on these cells in a dose-dependent manner. In addition, the plasma membranes of different cell lines and tissues from normal and autoimmune mice were isolated and probed by anti-dsDNA antibodies in Western blot analysis. We found the actively proliferating cells such as MC, RBA-1 and 3T3 may express both P0 (38,000 MW) and P1 (19,000 MW) on the surface membrane. In addition, the kidney, liver and spleen from either autoimmune MRL-lpr/lpr or BALB/c mice may constantly express P0 protein, but the expression of P1 is inconsistent. In contrast, brain and muscle from either mice failed to express P proteins on their surface. Unexpectedly, a high molecular weight substance (larger than 205,000 MW) with unknown nature appears in the membrane of brain and muscle tissues in both mice. Immunoprecipitation of the surface-biotinylated MC-lysate by anti-dsDNA antibodies further confirmed that P1 (19,000 MW) and P2 (17,000 MW) are really expressed on the cell surface. These results suggest that P proteins expressed on the surface of different tissues become the targets for anti-dsDNA antibodies mediating pleomorphic tissue

  2. A comparison of error detection rates between the reading aloud method and the double data entry method.

    PubMed

    Kawado, Miyuki; Hinotsu, Shiro; Matsuyama, Yutaka; Yamaguchi, Takuhiro; Hashimoto, Shuji; Ohashi, Yasuo

    2003-10-01

    Data entry and its verification are important steps in the process of data management in clinical studies. In Japan, a kind of visual comparison called the reading aloud (RA) method is often used as an alternative to or in addition to the double data entry (DDE) method. In a typical RA method, one operator reads previously keyed data aloud while looking at a printed sheet or computer screen, and another operator compares the voice with the corresponding data recorded on case report forms (CRFs) to confirm whether the data are the same. We compared the efficiency of the RA method with that of the DDE method in the data management system of the Japanese Registry of Renal Transplantation. Efficiency was evaluated in terms of error detection rate and expended time. Five hundred sixty CRFs were randomly allocated to two operators for single data entry. Two types of DDE and RA methods were performed. Single data entry errors were detected in 358 of 104,720 fields (per-field error rate=0.34%). Error detection rates were 88.3% for the DDE method performed by a different operator, 69.0% for the DDE method performed by the same operator, 59.5% for the RA method performed by a different operator, and 39.9% for the RA method performed by the same operator. The differences in these rates were significant (p<0.001) between the two verification methods as well as between the types of operator (same or different). The total expended times were 74.8 hours for the DDE method and 57.9 hours for the RA method. These results suggest that in detecting errors of single data entry, the RA method is inferior to the DDE method, while its time cost is lower. PMID:14500053

  3. Double hexagonal graphene ring synthesized using a growth-etching method.

    PubMed

    Liu, Jinyang; Xu, Yangyang; Cai, Hongbing; Zuo, Chuandong; Huang, Zhigao; Lin, Limei; Guo, Xiaomin; Chen, Zhendong; Lai, Fachun

    2016-08-01

    Precisely controlling the layer number, stacking order, edge configuration, shape and structure of graphene is extremely challenging but highly desirable in scientific research. In this report, a new concept named the growth-etching method has been explored to synthesize a graphene ring using the chemical vapor deposition process. The graphene ring is a hexagonal structure, which contains a hexagonal exterior edge and a hexagonal hole in the centre region. The most important concept introduced here is that the oxide nanoparticle derived from annealing is found to play a dual role. Firstly, it acts as a nucleation site to grow the hexagonal graphene domain and then it works as a defect for etching to form a hole. The evolution process of the graphene ring with the etching time was carefully studied. In addition, a double hexagonal graphene ring was successfully synthesized for the first time by repeating the growth-etching process, which not only confirms the validity and repeatability of the method developed here but may also be further extended to grow unique graphene nanostructures with three, four, or even tens of graphene rings. Finally, a schematic model was drawn to illustrate how the double hexagonal graphene ring is generated and propagated. The results shown here may provide valuable guidance for the design and growth of unique nanostructures of graphene and other two-dimensional materials. PMID:27387556

  4. Double hexagonal graphene ring synthesized using a growth-etching method

    NASA Astrophysics Data System (ADS)

    Liu, Jinyang; Xu, Yangyang; Cai, Hongbing; Zuo, Chuandong; Huang, Zhigao; Lin, Limei; Guo, Xiaomin; Chen, Zhendong; Lai, Fachun

    2016-07-01

    Precisely controlling the layer number, stacking order, edge configuration, shape and structure of graphene is extremely challenging but highly desirable in scientific research. In this report, a new concept named the growth-etching method has been explored to synthesize a graphene ring using the chemical vapor deposition process. The graphene ring is a hexagonal structure, which contains a hexagonal exterior edge and a hexagonal hole in the centre region. The most important concept introduced here is that the oxide nanoparticle derived from annealing is found to play a dual role. Firstly, it acts as a nucleation site to grow the hexagonal graphene domain and then it works as a defect for etching to form a hole. The evolution process of the graphene ring with the etching time was carefully studied. In addition, a double hexagonal graphene ring was successfully synthesized for the first time by repeating the growth-etching process, which not only confirms the validity and repeatability of the method developed here but may also be further extended to grow unique graphene nanostructures with three, four, or even tens of graphene rings. Finally, a schematic model was drawn to illustrate how the double hexagonal graphene ring is generated and propagated. The results shown here may provide valuable guidance for the design and growth of unique nanostructures of graphene and other two-dimensional materials.

  5. Trans-splicing as a novel method to rapidly produce antibody fusion proteins

    SciTech Connect

    Iwasaki, Ryohei; Kiuchi, Hiroki; Ihara, Masaki; Mori, Toshihiro; Kawakami, Masayuki; Ueda, Hiroshi

    2009-07-03

    To cultivate the use of trans-splicing as a novel means to rapidly express various antibody fusion proteins, we tried to express antibody-reporter enzyme fusions in a COS-1 co-transfection model. When a vector designed to induce trans-splicing with IgH pre-mRNA was co-transfected with a vector encoding the mouse IgM locus, the expression of V{sub H}-secreted human placental alkaline phosphatase (SEAP) as well as Fab-SEAP were successfully expressed both in mRNA and protein levels. Especially, the vectors encoding complementary sequence to S{mu} as a binding domain was accurate and efficient, producing trans-spliced mRNA of up to 2% of cis-spliced one. Since S{mu} sequence should exist in every IgH pre-mRNA, our finding will lead to the rapid production and analysis of various antibody-enzyme fusions suitable for enzyme-linked immunosorbent assay (ELISA) or antibody-dependent enzyme prodrug therapy (ADEPT).

  6. A double-observer method to estimate detection rate during aerial waterfowl surveys

    USGS Publications Warehouse

    Koneff, M.D.; Royle, J. Andrew; Otto, M.C.; Wortham, J.S.; Bidwell, J.K.

    2008-01-01

    We evaluated double-observer methods for aerial surveys as a means to adjust counts of waterfowl for incomplete detection. We conducted our study in eastern Canada and the northeast United States utilizing 3 aerial-survey crews flying 3 different types of fixed-wing aircraft. We reconciled counts of front- and rear-seat observers immediately following an observation by the rear-seat observer (i.e., on-the-fly reconciliation). We evaluated 6 a priori models containing a combination of several factors thought to influence detection probability including observer, seat position, aircraft type, and group size. We analyzed data for American black ducks (Anas rubripes) and mallards (A. platyrhynchos), which are among the most abundant duck species in this region. The best-supported model for both black ducks and mallards included observer effects. Sample sizes of black ducks were sufficient to estimate observer-specific detection rates for each crew. Estimated detection rates for black ducks were 0.62 (SE = 0.10), 0.63 (SE = 0.06), and 0.74 (SE = 0.07) for pilot-observers, 0.61 (SE = 0.08), 0.62 (SE = 0.06), and 0.81 (SE = 0.07) for other front-seat observers, and 0.43 (SE = 0.05), 0.58 (SE = 0.06), and 0.73 (SE = 0.04) for rear-seat observers. For mallards, sample sizes were adequate to generate stable maximum-likelihood estimates of observer-specific detection rates for only one aerial crew. Estimated observer-specific detection rates for that crew were 0.84 (SE = 0.04) for the pilot-observer, 0.74 (SE = 0.05) for the other front-seat observer, and 0.47 (SE = 0.03) for the rear-seat observer. Estimated observer detection rates were confounded by the position of the seat occupied by an observer, because observers did not switch seats, and by land-cover because vegetation and landform varied among crew areas. Double-observer methods with on-the-fly reconciliation, although not without challenges, offer one viable option to account for detection bias in aerial waterfowl

  7. Progress in the development of immunoanalytical methods incorporating recombinant antibodies to small molecular weight biotoxins.

    PubMed

    Kavanagh, Owen; Elliott, Christopher T; Campbell, Katrina

    2015-04-01

    Rapid immunoanalytical screening of food and environmental samples for small molecular weight (hapten) biotoxin contaminations requires the production of antibody reagents that possess the requisite sensitivity and specificity. To date animal-derived polyclonal (pAb) and monoclonal (mAb) antibodies have provided the binding element of the majority of these assays but recombinant antibodies (rAb) isolated from in vitro combinatorial phage display libraries are an exciting alternative due to (1) circumventing the need for experimental animals, (2) speed of production in commonly used in vitro expression systems and (3) subsequent molecular enhancement of binder performance. Short chain variable fragments (scFv) have been the most commonly employed rAb reagents for hapten biotoxin detection over the last two decades but antibody binding fragments (Fab) and single domain antibodies (sdAb) are increasing in popularity due to increased expression efficiency of functional binders and superior resistance to solvents. rAb-based immunochromatographic assays and surface plasmon resonance (SPR) biosensors have been reported to detect sub-regulatory levels of fungal (mycotoxins), marine (phycotoxins) and aquatic biotoxins in a wide range of food and environmental matrices, however this technology has yet to surpass the performances of the equivalent mAb- and pAb-based formats. As such the full potential of rAb technology in hapten biotoxin detection has yet to be achieved, but in time the inherent advantages of engineered rAb are set to provide the next generation of ultra-high performing binder reagents for the rapid and specific detection of hapten biotoxins. PMID:25716465

  8. Research on absorption test methods of Yb-doped double cladding fiber

    NASA Astrophysics Data System (ADS)

    Wang, Pupu; Li, Rundong; Rong, Liang; Ji, Wei; Gao, Yankun; Jiang, Cong; Gu, Shaoyi

    2016-01-01

    Absorption coefficient is a very useful feature for active fiber. In fiber laser system, the length of active fiber is chosen according to absorption coefficient. And the length of fiber can directly influence the feature of fiber laser. Therefore, how to obtain an accurate absorption coefficient is very important. Because fiber exists re-emission in typical absorption band pumped by power. It is difficult to accurately measure absorption coefficient. The absorption coefficients of Yb-doped double cladding fiber at 975 nm measured by several methods were compared. In conclusion, for the fibers with same length pumped by white light, the absorption coefficient is the highest when cutback only once. Meanwhile, when fibers with different length were measured by the same method, the absorption coefficient is inversely proportional to optical fiber length.

  9. Simulation of electric double-layer capacitors: evaluation of constant potential method

    NASA Astrophysics Data System (ADS)

    Wang, Zhenxing; Laird, Brian; Yang, Yang; Olmsted, David; Asta, Mark

    2014-03-01

    Atomistic simulations can play an important role in understanding electric double-layer capacitors (EDLCs) at a molecular level. In such simulations, typically the electrode surface is modeled using fixed surface charges, which ignores the charge fluctuation induced by local fluctuations in the electrolyte solution. In this work we evaluate an explicit treatment of charges, namely constant potential method (CPM)[1], in which the electrode charges are dynamically updated to maintain constant electrode potential. We employ a model system with a graphite electrode and a LiClO4/acetonitrile electrolyte, examined as a function of electrode potential differences. Using various molecular and macroscopic properties as metrics, we compare CPM simulations on this system to results using fixed surface charges. Specifically, results for predicted capacity, electric potential gradient and solvent density profile are identical between the two methods; However, ion density profiles and solvation structure yield significantly different results.

  10. Method for determining effective nonradiative lifetime and leakage losses in double-heterostructure lasers

    SciTech Connect

    van Opdorp, C.; 't Hooft, G.W.

    1981-06-01

    Carrier losses in double-heterostructure lasers are twofold: (i) nonradiative recombination through killers in the bulk of the active region and at all its boundaries (interfaces and surfaces), and (ii) leakage out of the active region. A simple theory shows the following. In the high-injection regime (papprox. =n) all processes under (i) are directly proportional to n. Consequently their contributions can be lumped together in a single effective nonradiative carrier lifetime tau/sub nr/ ; this tau/sub nr/ is constant (i.e., independent of n) owing to the constant degree of occupation of all killers in the mentioned regime. On the other hand, the leakage losses (ii) are superlinear in n. This provides a well-grounded basis for disentangling the contributions of (i) and (ii) in a given sample. Further, a simple method is presented for accurately determining tau/sub nr/ from data of the external quantum efficiency eta/sub ext/ measured as a function of current I in the spontaneous high-injection regime below the laser threshold. Knowledge of the light-extraction factor (i.e., the ratio of external and internal quantum efficiencies) is essentially unnecessary with this method. However, optionally it can be determined easily from a slight extension of the method. For illustration the method of determining tau/sub nr/, which is also applicable to double-hetero LED's, has been applied to some thirty LPE and metal-organic VPE GaAs-(Ga,Al)As lasers of widely varying qualities. The values found vary between 0.8 and 55 ns. From the measured values of tau/sub nr/ it follows that the upper limit for the interface recombination velocity in the best samples is 270 cm/s. For most samples tau/sub nr/ cannot account for all electrical losses at laser threshold. The superlinear excess losses are ascribable to leakage.

  11. A new three-dimensional shape measurement method based on double-frequency fringes

    NASA Astrophysics Data System (ADS)

    Li, Biao; Yang, Jie; Wu, Haitao; Fu, Yanjun

    2015-10-01

    Fringe projection profilometry (FPP) is a rapidly developing technique which is widely used for industrial manufacture, heritage conservation, and medicine etc. because of its high speed, high precision, non-contact operation, full-field acquisition, and easy information processing. Among the various FFP methods, the squared binary defocused projection method (SBM) has been promptly expanding with several advantages: (1) high projection speed because of 1-bit grayscale fringe; (2) eliminating nonlinear gamma of the projector for the defocusing effect. Nevertheless, the method is not trouble-free. When the fringe stripe is wide, it brings down the fringe contrast and is difficult to control the defocused degree, resulting in a low measurement accuracy. In order to further improve high-speed and high-precision three-dimensional shape measurement, this paper presents a new three-dimensional shape measurement method based on double-frequency fringes projection. This new method needs to project two sets of 1-bit grayscale fringe patterns (low-frequency fringe and high-frequency fringe) onto the object surface under slightly defocused projection mode. The method has the following advantages: (1) high projection speed because of 1-bit grayscale fringe; (2) high measurement precision for selectively removing undesired harmonics. Low-frequency fringe is produced by error-diffusion dithering (Dithering) technique and high-frequency fringe is generated by optimal pulse-width modulation (OPWM) technique. The two kinds of fringe patterns have each superiorities and flaws. The low-frequency fringe has a low measurement accuracy, but the continue phase can be easily retrieved. However, the property of high-frequency fringe and low-frequency fringe is the opposite. The general idea of this method proposed is as follows: Because the both fringes test the same object, the height is the same. The low-frequency fringe can be used to assist the high frequency fringe to retrieve

  12. Enzyme-labeled Antigen Method: Development and Application of the Novel Approach for Identifying Plasma Cells Locally Producing Disease-specific Antibodies in Inflammatory Lesions

    PubMed Central

    Mizutani, Yasuyoshi; Shiogama, Kazuya; Onouchi, Takanori; Sakurai, Kouhei; Inada, Ken-ichi; Tsutsumi, Yutaka

    2016-01-01

    In chronic inflammatory lesions of autoimmune and infectious diseases, plasma cells are frequently observed. Antigens recognized by antibodies produced by the plasma cells mostly remain unclear. A new technique identifying these corresponding antigens may give us a breakthrough for understanding the disease from a pathophysiological viewpoint, simply because the immunocytes are seen within the lesion. We have developed an enzyme-labeled antigen method for microscopic identification of the antigen recognized by specific antibodies locally produced in plasma cells in inflammatory lesions. Firstly, target biotinylated antigens were constructed by the wheat germ cell-free protein synthesis system or through chemical biotinylation. Next, proteins reactive to antibodies in tissue extracts were screened and antibody titers were evaluated by the AlphaScreen method. Finally, with the enzyme-labeled antigen method using the biotinylated antigens as probes, plasma cells producing specific antibodies were microscopically localized in fixed frozen sections. Our novel approach visualized tissue plasma cells that produced 1) autoantibodies in rheumatoid arthritis, 2) antibodies against major antigens of Porphyromonas gingivalis in periodontitis or radicular cyst, and 3) antibodies against a carbohydrate antigen, Strep A, of Streptococcus pyogenes in recurrent tonsillitis. Evaluation of local specific antibody responses expectedly contributes to clarifying previously unknown processes in inflammatory disorders. PMID:27006517

  13. MARCC (Matrix-Assisted Reader Chromatin Capture): an antibody-free method to enrich and analyze combinatorial nucleosome modifications

    PubMed Central

    Su, Zhangli

    2016-01-01

    Combinatorial patterns of histone modifications are key indicators of different chromatin states. Most of the current approaches rely on the usage of antibodies to analyze combinatorial histone modifications. Here we detail an antibody-free method named MARCC (Matrix-Assisted Reader Chromatin Capture) to enrich combinatorial histone modifications. The combinatorial patterns are enriched on native nucleosomes extracted from cultured mammalian cells and prepared by micrococcal nuclease digestion. Such enrichment is achieved by recombinant chromatin-interacting protein modules, or so-called reader domains, which can bind in a combinatorial modification-dependent manner. The enriched chromatin can be quantified by western blotting or mass spectrometry for the co-existence of histone modifications, while the associated DNA content can be analyzed by qPCR or next-generation sequencing. Altogether, MARCC provides a reproducible, efficient and customizable solution to enrich and analyze combinatorial histone modifications. PMID:26131849

  14. Double isotopic method using dansyl chloride for the determination of GABA in rat C6 astrocytoma cell cultures

    SciTech Connect

    Kohl, R.L.; Quay, W.B.; Perez-Polo, J.R.

    1986-01-01

    Methods are described for the quantitative measurement of GABA in culture. The method can be adapted to any amino acid or dansyl-chloride-reactive species. The sensitivity and selectivity of the procedure result from the double isotopic design in which (/sup 14/C)-labeled internal standard was added to the samples before reaction with (3M)-labeled dansyl chloride. Values obtained by ion-exchange amino acid analysis of cultures agree closely with the values obtained by the double isotopic method. This method is sensitive enough to measure GABA intracellularly and the condition medium.

  15. SEC Based Method for Size Determination of Immune Complexes of Therapeutic Antibodies in Animal Matrix

    PubMed Central

    Boysen, Marta; Dreher, Ingeborg; Loebbert, Ralf

    2016-01-01

    Therapeutic monoclonal antibodies (mAbs) represent a milestone in pharmacological development. Their superiority is based on the combination of high specificity, low toxicity, and long half-life that characterizes biologics. If biologics have Achilles' heel, it is their potential immunogenicity. To better understand the impact of the size of immune complexes of mAbs on anti-drug antibody (ADA) dependent adverse reactions in Macaca fascicularis, we developed an efficient high-throughput size exclusion chromatography- (SEC-) based methodology that enables analysis of the size, size distribution, and ratio of free and ADA-complexed mAb in serum allowing for assessment of formation and clearance of circulating ADA-mAb immune complexes (CIC). PMID:27556050

  16. Equation-of-motion coupled cluster method for high spin double electron attachment calculations

    SciTech Connect

    Musiał, Monika Lupa, Łukasz; Kucharski, Stanisław A.

    2014-03-21

    The new formulation of the equation-of-motion (EOM) coupled cluster (CC) approach applicable to the calculations of the double electron attachment (DEA) states for the high spin components is proposed. The new EOM equations are derived for the high spin triplet and quintet states. In both cases the new equations are easier to solve but the substantial simplification is observed in the case of quintets. Out of 21 diagrammatic terms contributing to the standard DEA-EOM-CCSDT equations for the R{sub 2} and R{sub 3} amplitudes only four terms survive contributing to the R{sub 3} part. The implemented method has been applied to the calculations of the excited states (singlets, triplets, and quintets) energies of the carbon and silicon atoms and potential energy curves for selected states of the Na{sub 2} (triplets) and B{sub 2} (quintets) molecules.

  17. Simulated likelihood methods for complex double-platform line transect surveys.

    PubMed

    Schweder, T; Skaug, H J; Langaas, M; Dimakos, X K

    1999-09-01

    The conventional line transect approach of estimating effective search width from the perpendicular distance distribution is inappropriate in certain types of surveys, e.g., when an unknown fraction of the animals on the track line is detected, the animals can be observed only at discrete points in time, there are errors in positional measurements, and covariate heterogeneity exists in detectability. For such situations a hazard probability framework for independent observer surveys is developed. The likelihood of the data, including observed positions of both initial and subsequent observations of animals, is established under the assumption of no measurement errors. To account for measurement errors and possibly other complexities, this likelihood is modified by a function estimated from extensive simulations. This general method of simulated likelihood is explained and the methodology applied to data from a double-platform survey of minke whales in the northeastern Atlantic in 1995. PMID:11314993

  18. Study of acoustic field modulation in the regenerator by double loudspeakers method.

    PubMed

    Zhou, Lihua; Xie, Xiujuan; Li, Qing

    2011-11-01

    A model to modulate acoustic field in a regenerator of a thermoacoustic system by the double loudspeakers method is presented in this paper. The equations are derived for acoustic field modulation. They represent the relations among acoustic field (complex pressure p(0), complex velocity u(0), and acoustic impedance Z(0)), driving parameters of loudspeakers (voltage amplitude and its phase difference), and operating parameters involved in a matrix H (frequency, temperature of regenerator). The range of acoustic field is adjustable and limited by the maximal driving voltages of loudspeakers according to driving parameters. The range is simulated and analyzed in the amplitude-phase and complex coordinate planes for a given or variable H. The simulated results indicate that the range has its intrinsic characteristics. The expected acoustic field in a regenerator can be obtained feasibly by the modulation. PMID:22087899

  19. 1-D seismic velocity model and hypocenter relocation using double difference method around West Papua region

    SciTech Connect

    Sabtaji, Agung E-mail: agung.sabtaji@bmkg.go.id; Nugraha, Andri Dian

    2015-04-24

    West Papua region has fairly high of seismicity activities due to tectonic setting and many inland faults. In addition, the region has a unique and complex tectonic conditions and this situation lead to high potency of seismic hazard in the region. The precise earthquake hypocenter location is very important, which could provide high quality of earthquake parameter information and the subsurface structure in this region to the society. We conducted 1-D P-wave velocity using earthquake data catalog from BMKG for April, 2009 up to March, 2014 around West Papua region. The obtained 1-D seismic velocity then was used as input for improving hypocenter location using double-difference method. The relocated hypocenter location shows fairly clearly the pattern of intraslab earthquake beneath New Guinea Trench (NGT). The relocated hypocenters related to the inland fault are also observed more focus in location around the fault.

  20. A double-observer method for reducing bias in faecal pellet surveys of forest ungulates

    USGS Publications Warehouse

    Jenkins, K.J.; Manly, B.F.J.

    2008-01-01

    1. Faecal surveys are used widely to study variations in abundance and distribution of forest-dwelling mammals when direct enumeration is not feasible. The utility of faecal indices of abundance is limited, however, by observational bias and variation in faecal disappearance rates that obscure their relationship to population size. We developed methods to reduce variability in faecal surveys and improve reliability of faecal indices. 2. We used double-observer transect sampling to estimate observational bias of faecal surveys of Roosevelt elk Cervus elaphus roosevelti and Columbian black-tailed deer Odocoileus hemionus columbianus in Olympic National Park, Washington, USA. We also modelled differences in counts of faecal groups obtained from paired cleared and uncleared transect segments as a means to adjust standing crop faecal counts for a standard accumulation interval and to reduce bias resulting from variable decay rates. 3. Estimated detection probabilities of faecal groups ranged from < 0.2-1.0 depending upon the observer, whether the faecal group was from elk or deer, faecal group size, distance of the faecal group from the sampling transect, ground vegetation cover, and the interaction between faecal group size and distance from the transect. 4. Models of plot-clearing effects indicated that standing crop counts of deer faecal groups required 34% reduction on flat terrain and 53% reduction on sloping terrain to represent faeces accumulated over a standard 100-day interval, whereas counts of elk faecal groups required 0% and 46% reductions on flat and sloping terrain, respectively. 5. Synthesis and applications. Double-observer transect sampling provides a cost-effective means of reducing observational bias and variation in faecal decay rates that obscure the interpretation of faecal indices of large mammal abundance. Given the variation we observed in observational bias of faecal surveys and persistence of faeces, we emphasize the need for future

  1. ISS Double-Gimbaled CMG Subsystem Simulation Using the Agile Development Method

    NASA Technical Reports Server (NTRS)

    Inampudi, Ravi

    2016-01-01

    This paper presents an evolutionary approach in simulating a cluster of 4 Control Moment Gyros (CMG) on the International Space Station (ISS) using a common sense approach (the agile development method) for concurrent mathematical modeling and simulation of the CMG subsystem. This simulation is part of Training systems for the 21st Century simulator which will provide training for crew members, instructors, and flight controllers. The basic idea of how the CMGs on the space station are used for its non-propulsive attitude control is briefly explained to set up the context for simulating a CMG subsystem. Next different reference frames and the detailed equations of motion (EOM) for multiple double-gimbal variable-speed control moment gyroscopes (DGVs) are presented. Fixing some of the terms in the EOM becomes the special case EOM for ISS's double-gimbaled fixed speed CMGs. CMG simulation development using the agile development method is presented in which customer's requirements and solutions evolve through iterative analysis, design, coding, unit testing and acceptance testing. At the end of the iteration a set of features implemented in that iteration are demonstrated to the flight controllers thus creating a short feedback loop and helping in creating adaptive development cycles. The unified modeling language (UML) tool is used in illustrating the user stories, class designs and sequence diagrams. This incremental development approach of mathematical modeling and simulating the CMG subsystem involved the development team and the customer early on, thus improving the quality of the working CMG system in each iteration and helping the team to accurately predict the cost, schedule and delivery of the software.

  2. Uniscale multi-view registration using double dog-leg method

    NASA Astrophysics Data System (ADS)

    Chen, Chao-I.; Sargent, Dusty; Tsai, Chang-Ming; Wang, Yuan-Fang; Koppel, Dan

    2009-02-01

    3D computer models of body anatomy can have many uses in medical research and clinical practices. This paper describes a robust method that uses videos of body anatomy to construct multiple, partial 3D structures and then fuse them to form a larger, more complete computer model using the structure-from-motion framework. We employ the Double Dog-Leg (DDL) method, a trust-region based nonlinear optimization method, to jointly optimize the camera motion parameters (rotation and translation) and determine a global scale that all partial 3D structures should agree upon. These optimized motion parameters are used for constructing local structures, and the global scale is essential for multi-view registration after all these partial structures are built. In order to provide a good initial guess of the camera movement parameters and outlier free 2D point correspondences for DDL, we also propose a two-stage scheme where multi-RANSAC with a normalized eight-point algorithm is first performed and then a few iterations of an over-determined five-point algorithm is used to polish the results. Our experimental results using colonoscopy video show that the proposed scheme always produces more accurate outputs than the standard RANSAC scheme. Furthermore, since we have obtained many reliable point correspondences, time-consuming and error-prone registration methods like the iterative closest points (ICP) based algorithms can be replaced by a simple rigid-body transformation solver when merging partial structures into a larger model.

  3. Comparing high-throughput methods to measure NK cell-mediated antibody dependent cellular cytotoxicity during HIV-infection.

    PubMed

    Konstantinus, Iyaloo N; Gamieldien, Hoyam; Mkhize, Nonhlanhla N; Kriek, Jean-Mari; Passmore, Jo-Ann S

    2016-07-01

    HIV-specific binding antibody responses, including those mediating antibody-dependent cellular cytotoxicity (ADCC), provided the best functional correlate of lower risk of infection in the RV144 HIV-1 vaccine clinical trial. The aim of this study was to compare two high-throughput flow cytometry based methods to measure HIV-specific ADCC responses, the GranToxilux and PanToxilux assays. Plasma from nine HIV-1 seropositive individuals was screened for binding antibody titres against HIV-1 subtype C gp120 by ELISA and western blot. Plasma from six HIV-negative individuals was included as controls. Both ADCC assays used subtype C gp120-coated CEM.NKRCCR5 cells as targets. The PanToxilux assay (which measured both granzyme B and caspase activity) measured higher levels of direct natural killer (NK) cell killing of K562 tumour cells than the GranToxilux assay (granzyme B alone; p<0.05). In ADCC assays in which NK cell killing was directed against gp120-coated CEM.NKRCCR5 cells in an antibody-dependent manner, plasma from HIV-positive individuals yielded significantly higher levels of ADCC activity than the HIV-negative controls. In contrast to direct killing, the GranToxilux assay measured similar levels of ADCC killing as the PanToxilux assay but had significantly lower background cytotoxicity against target cells coated with HIV negative serum. In conclusion, the PanToxilux assay was more sensitive for detecting direct NK cell killing of K562 cells than the GranToxilux assay, although the GranToxilux assay performed better at detecting HIV-specific ADCC activity, because of lower background cytotoxicity from HIV-negative serum. This is the first study to compare GranToxilux and PanToxilux ability to detect ADCC during HIV infection. PMID:27094485

  4. Validation of an automated method for compounding monoclonal antibody patient doses

    PubMed Central

    Peters, Bas J.M.; Capelle, Martinus A.H.; Arvinte, Tudor; van de Garde, Ewoudt M.W.

    2013-01-01

    Automation robots have recently come to the market as an alternative for manual compounding of drugs for intravenous administration. Our aim was to assess whether robotic compounding can be performed with monoclonal antibodies (mAbs) without influencing the aggregation state of the proteins. Three frequently used mAbs were studied: infliximab (Remicade®, Janssen Biotech) and trastuzumab (Herceptin®, Roche) in lyophilised form, and bevacizumab (Avastin®, Roche) as a liquid formulation stored at 2°C to 8°C. The effects of different procedures to prepare the patient doses on antibody aggregation were evaluated. Remicade® and Herceptin® were reconstituted both manually and by a robotic arm (i.v.STATION®, Health Robotics). Additionally, the influence of vigorous shaking during reconstitution was investigated. The effects of rapid aspiration and dispensing on antibody aggregation were investigated for all three mAbs. Aggregation state was assessed by UV-Vis absorbance, 90° light scatter, fluorescence spectroscopy, Nile red fluorescence microscopy, and field flow fractionation without cross and focus flow. Robotic reconstituted samples showed similar findings compared with manual reconstitution if performed exactly according to the summary of product characteristics (SPC). Vials that were vigorously shaken showed a significant increase in aggregates. Similarly, rapid aspiration/dispense cycles resulted in a strong increase in the number and sizes of aggregates for all three mAbs; this result was observed after just one rapid aspiration/dispense cycle. Our study showed that robotic compounding of mAbs is feasible if the robot is exactly programmed according to the SPC, indicating that robotic compounding can be used to achieve reproducible high-quality compounding for delicate formulations. PMID:23255057

  5. Enrollment Forecasting with Double Exponential Smoothing: Two Methods for Objective Weight Factor Selection. AIR Forum 1980 Paper.

    ERIC Educational Resources Information Center

    Gardner, Don E.

    The merits of double exponential smoothing are discussed relative to other types of pattern-based enrollment forecasting methods. The difficulties associated with selecting an appropriate weight factor are discussed, and their potential effects on prediction results are illustrated. Two methods for objectively selecting the "best" weight factor…

  6. Evaluation of the constant potential method in simulating electric double-layer capacitors.

    PubMed

    Wang, Zhenxing; Yang, Yang; Olmsted, David L; Asta, Mark; Laird, Brian B

    2014-11-14

    A major challenge in the molecular simulation of electric double layer capacitors (EDLCs) is the choice of an appropriate model for the electrode. Typically, in such simulations the electrode surface is modeled using a uniform fixed charge on each of the electrode atoms, which ignores the electrode response to local charge fluctuations in the electrolyte solution. In this work, we evaluate and compare this Fixed Charge Method (FCM) with the more realistic Constant Potential Method (CPM), [S. K. Reed et al., J. Chem. Phys. 126, 084704 (2007)], in which the electrode charges fluctuate in order to maintain constant electric potential in each electrode. For this comparison, we utilize a simplified LiClO4-acetonitrile/graphite EDLC. At low potential difference (ΔΨ ⩽ 2 V), the two methods yield essentially identical results for ion and solvent density profiles; however, significant differences appear at higher ΔΨ. At ΔΨ ⩾ 4 V, the CPM ion density profiles show significant enhancement (over FCM) of "inner-sphere adsorbed" Li(+) ions very close to the electrode surface. The ability of the CPM electrode to respond to local charge fluctuations in the electrolyte is seen to significantly lower the energy (and barrier) for the approach of Li(+) ions to the electrode surface. PMID:25399127

  7. A hybrid ensemble method based on double disturbance for classifying microarray data.

    PubMed

    Chen, Tao; Xue, Huifeng; Hong, Zenglin; Cui, Man; Zhao, Hui

    2015-01-01

    Microarray data has small samples and high dimension, and it contains a significant amount of irrelevant and redundant genes. This paper proposes a hybrid ensemble method based on double disturbance to improve classification performance. Firstly, original genes are ranked through reliefF algorithm and part of the genes are selected from the original genes set, and then a new training set is generated from the original training set according to the previously selected genes. Secondly, D bootstrap training subsets are produced from the previously generated training set by bootstrap technology. Thirdly, an attribute reduction method based on neighborhood mutual information with a different radius is used to reduce genes on each bootstrap training subset to produce new training subsets. Each new training subset is applied to train a base classifier. Finally, a part of the base classifiers are selected based on the teaching-learning-based optimization to build an ensemble by weighted voting. Experimental results on six benchmark cancer microarray datasets showed proposed method decreased ensemble size and obtained higher classification performance compared with Bagging, AdaBoost, and Random Forest. PMID:26405970

  8. Low-order mathematical modelling of electric double layer supercapacitors using spectral methods

    NASA Astrophysics Data System (ADS)

    Drummond, Ross; Howey, David A.; Duncan, Stephen R.

    2015-03-01

    This work investigates two physics-based models that simulate the non-linear partial differential algebraic equations describing an electric double layer supercapacitor. In one model the linear dependence between electrolyte concentration and conductivity is accounted for, while in the other model it is not. A spectral element method is used to discretise the model equations and it is found that the error convergence rate with respect to the number of elements is faster compared to a finite difference method. The increased accuracy of the spectral element approach means that, for a similar level of solution accuracy, the model simulation computing time is approximately 50% of that of the finite difference method. This suggests that the spectral element model could be used for control and state estimation purposes. For a typical supercapacitor charging profile, the numerical solutions from both models closely match experimental voltage and current data. However, when the electrolyte is dilute or where there is a long charging time, a noticeable difference between the numerical solutions of the two models is observed. Electrical impedance spectroscopy simulations show that the capacitance of the two models rapidly decreases when the frequency of the perturbation current exceeds an upper threshold.

  9. Evaluation of the constant potential method in simulating electric double-layer capacitors

    NASA Astrophysics Data System (ADS)

    Wang, Zhenxing; Yang, Yang; Olmsted, David L.; Asta, Mark; Laird, Brian B.

    2014-11-01

    A major challenge in the molecular simulation of electric double layer capacitors (EDLCs) is the choice of an appropriate model for the electrode. Typically, in such simulations the electrode surface is modeled using a uniform fixed charge on each of the electrode atoms, which ignores the electrode response to local charge fluctuations in the electrolyte solution. In this work, we evaluate and compare this Fixed Charge Method (FCM) with the more realistic Constant Potential Method (CPM), [S. K. Reed et al., J. Chem. Phys. 126, 084704 (2007)], in which the electrode charges fluctuate in order to maintain constant electric potential in each electrode. For this comparison, we utilize a simplified LiClO4-acetonitrile/graphite EDLC. At low potential difference (ΔΨ ⩽ 2 V), the two methods yield essentially identical results for ion and solvent density profiles; however, significant differences appear at higher ΔΨ. At ΔΨ ⩾ 4 V, the CPM ion density profiles show significant enhancement (over FCM) of "inner-sphere adsorbed" Li+ ions very close to the electrode surface. The ability of the CPM electrode to respond to local charge fluctuations in the electrolyte is seen to significantly lower the energy (and barrier) for the approach of Li+ ions to the electrode surface.

  10. Evaluation of the constant potential method in simulating electric double-layer capacitors

    SciTech Connect

    Wang, Zhenxing; Laird, Brian B.; Yang, Yang; Olmsted, David L.; Asta, Mark

    2014-11-14

    A major challenge in the molecular simulation of electric double layer capacitors (EDLCs) is the choice of an appropriate model for the electrode. Typically, in such simulations the electrode surface is modeled using a uniform fixed charge on each of the electrode atoms, which ignores the electrode response to local charge fluctuations in the electrolyte solution. In this work, we evaluate and compare this Fixed Charge Method (FCM) with the more realistic Constant Potential Method (CPM), [S. K. Reed et al., J. Chem. Phys. 126, 084704 (2007)], in which the electrode charges fluctuate in order to maintain constant electric potential in each electrode. For this comparison, we utilize a simplified LiClO{sub 4}-acetonitrile/graphite EDLC. At low potential difference (ΔΨ ⩽ 2 V), the two methods yield essentially identical results for ion and solvent density profiles; however, significant differences appear at higher ΔΨ. At ΔΨ ⩾ 4 V, the CPM ion density profiles show significant enhancement (over FCM) of “inner-sphere adsorbed” Li{sup +} ions very close to the electrode surface. The ability of the CPM electrode to respond to local charge fluctuations in the electrolyte is seen to significantly lower the energy (and barrier) for the approach of Li{sup +} ions to the electrode surface.

  11. Development of an antibody-based diagnostic method for the identification of Bemisia tabaci biotype B.

    PubMed

    Baek, Ji Hyeong; Lee, Hye Jung; Kim, Young Ho; Lim, Kook Jin; Lee, Si Hyeock; Kim, Bum Joon

    2016-07-01

    The whitefly Bemisia tabaci is a very destructive pest. B. tabaci is composed of various morphologically undistinguishable biotypes, among which biotypes B and Q, in particular, draw attention because of their wide distribution in Korea and differential potentials for insecticide resistance development. To develop a biotype-specific protein marker that can readily distinguishes biotypes B from other biotypes in the field, we established an ELISA protocol based on carboxylesterase 2 (COE2), which is more abundantly expressed in biotypes B compared with Q. Recombinant COE2 was expressed, purified and used for antibody construction. Polyclonal antibodies specific to B. tabaci COE2 [anti-COE2 pAb and deglycosylated anti-COE2 pAb (DG anti-COE2 pAb)] revealed a 3-9-fold higher reactivity to biotype B COE2 than biotype Q COE2 by Western blot and ELISA analyses. DG anti-COE2 pAb exhibited low non-specific activity, demonstrating its compatibility in diagnosing biotypes. Western blot and ELISA analyses determined that one of the 11 field populations examined was biotype B and the others were biotype Q, suggesting the saturation of biotype Q in Korea. DG anti-COE2 pAb discriminates B. tabaci biotypes B and Q with high specificity and accuracy and could be useful for the development of a B. tabaci biotype diagnosis kit for on-site field applications. PMID:27265822

  12. Multiplex method for initial complex testing of antibodies to blood transmitted diseases agents.

    PubMed

    Poltavchenko, Alexander G; Nechitaylo, Oleg V; Filatov, Pavel V; Ersh, Anna V; Gureyev, Vadim N

    2016-10-01

    Initial screening of donors and population at high risk of infection with blood transmitted diseases involves a number of analyses using monospesific diagnostic systems, and therefore is expensive labor- and time-consuming process. The goal of this work is to construct a multiplex test enabling to carry out rapid initial complex testing at a low price. The paper describes a kit making it possible to detect simultaneously antibodies to six agents of the most significant blood transmitted diseases: HIV virus, hepatitis B and C viruses, cytomegalovirus, T. pallidum and T. gondii in blood products. The kit comprises multiplex dot-immunoassay based on plane protein arrays (immune chips) using colloidal gold conjugates and silver development. It provides an opportunity to carry out complex analysis within 70min at room temperature, and there is no need of well-qualified personnel. We compared laboratory findings of the kit with monospecific kits for ELISA produced by two Russian commercial companies. Dot-assay results correlate well with data obtained using commercial kits for ELISA. Furthermore, multiplex analysis is quicker and cheaper in comparison with ELISA and can be carried out in non-laboratory conditions. The kit for multiplex dot-immunoassay of antibodies to blood transmitted agents can significantly simplify initial complex testing. PMID:27497868

  13. Calibration of Hydraulic Conductivities by the Kalman Filtered Double Constraint Method

    NASA Astrophysics Data System (ADS)

    Zijl, Wouter; El-Rawy, Mustafa; Batelaan, Okke

    2014-05-01

    To assess the consequences of a changing environment for future management decisions we need quantitative techniques validated by case studies. In this context dealing with the limited data availability and inherent uncertainty is a major challenge. In this contribution we present a combination of two techniques (the Double Constraint Method and the Kalman Filter) exemplified by case studies. The techniques assist in the calibration of hydraulic grid block conductivities as well as in finding the reliability of the result. To focus on the basic principles we exemplify our approach for flow in which storage by water compressibility and pore space deformation is negligible. Only storage by water table movements plays a role. Such conditions hold for most flow problems in the relatively shallow aquifer-aquitard systems of deltaic regions. In a forward problem the conductivity is specified in all grid blocks. In addition, on each point of the boundary and in each well only one type of boundary condition has to be specified: either head, or flux. Our approach is based on the principle that calibration of the initial conductivities is meaningful only if we can specify both head and flux at a number of boundary points or wells, including no-flux monitoring wells. In general a hydrogeological model is a "flux model," i.e., the model is as much as possible based on specified ("measured") fluxes through the boundaries and in the wells. An exception is Tóth's flow systems analysis where, instead of the usual recharge fluxes, heads are specified on the water table. This suggests building a second forward model, a "head model," that is as much as possible based on specified (measured) heads on the boundaries and in the wells. The initial conductivities are then updated by applying Darcy's law K = -q/(¶h/¶x) to the fluxes q obtained by the "flux model" and the head gradients ¶h/¶x obtained by the "head model." This so-called "double constraint method" (DCM) leads to a

  14. A blind image detection method for information hiding with double random-phase encoding

    NASA Astrophysics Data System (ADS)

    Sheng, Yuan; Xin, Zhou; Jian-guo, Chen; Yong-liang, Xiao; Qiang, Liu

    2009-07-01

    In this paper, a blind image detection method based on a statistical hypothesis test for information hiding with double random-phase encoding (DRPE) is proposed. This method aims to establish a quantitative criterion which is used to judge whether there is secret information embedded in the detected image. The main process can be described as follows: at the beginning, we decompose the detected gray-scale image into 8 bit planes considering it has 256 gray levels, and suppose that a secret image has been hidden in the detected image after it was encrypted by DRPE, thus the lower bit planes of the detected image exhibit strong randomness. Then, we divide the bit plane to be tested into many windows, and establish a statistical variable to measure the relativity between pixels in every window. Finally, judge whether the secret image exists in the detected image by operating the t test on all statistical variables. Numerical simulation shows that the accuracy is quite satisfactory, when we need to distinguish the images carrying secret information from a large amount of images.

  15. A NUMERICAL METHOD FOR STUDYING SUPER-EDDINGTON MASS TRANSFER IN DOUBLE WHITE DWARF BINARIES

    SciTech Connect

    Marcello, Dominic C.; Tohline, Joel E. E-mail: tohline@phys.lsu.edu

    2012-04-01

    We present a numerical method for the study of double white dwarf (DWD) binary systems at the onset of super-Eddington mass transfer. We incorporate the physics of ideal inviscid hydrodynamical flow, Newtonian self-gravity, and radiation transport on a three-dimensional uniformly rotating cylindrical Eulerian grid. Care has been taken to conserve the key physical quantities such as angular momentum and energy. Our new method conserves total energy to a higher degree of accuracy than other codes that are presently being used to model mass transfer in DWD systems. We present the results of verification tests and simulate the first 20 + orbits of a binary system of mass ratio q 0.7 at the onset of dynamically unstable direct impact mass transfer. The mass transfer rate quickly exceeds the critical Eddington limit by many orders of magnitude, and thus we are unable to model a trans-Eddington phase. It appears that radiation pressure does not significantly affect the accretion flow in the highly super-Eddington regime. An optically thick common envelope forms around the binary within a few orbits. Although this envelope quickly exceeds the spatial domain of the computational grid, the fraction of the common envelope that exceeds zero gravitational binding energy is extremely small, suggesting that radiation-driven mass loss is insignificant in this regime. It remains to be seen whether simulations that capture the trans-Eddington phase of such flows will lead to the same conclusion or show that substantial material gets expelled.

  16. Simple hydraulic conductivity estimation by the Kalman filtered double constraint method.

    PubMed

    El-Rawy, M A; Batelaan, O; Zijl, W

    2015-01-01

    This paper presents the Kalman Filtered Double Constraint Method (DCM-KF) as a technique to estimate the hydraulic conductivities in the grid blocks of a groundwater flow model. The DCM is based on two forward runs with the same initial grid block conductivities, but with alternating flux-head conditions specified on parts of the boundary and the wells. These two runs are defined as: (1) the flux run, with specified fluxes (recharge and well abstractions), and (2) the head run, with specified heads (measured in piezometers). Conductivities are then estimated as the initial conductivities multiplied by the fluxes obtained from the flux run and divided by the fluxes obtained from the head run. The DCM is easy to implement in combination with existing models (e.g., MODFLOW). Sufficiently accurate conductivities are obtained after a few iterations. Because of errors in the specified head-flux couples, repeated estimation under varying hydrological conditions results in different conductivities. A time-independent estimate of the conductivities and their inaccuracy can be obtained by a simple linear KF with modest computational requirements. For the Kleine Nete catchment, Belgium, the DCM-KF yields sufficiently accurate calibrated conductivities. The method also results in distinguishing regions where the head-flux observations influence the calibration from areas where it is not able to influence the hydraulic conductivity. PMID:24854328

  17. Double ionization of helium by fast electrons with the Generalized Sturmian Functions method

    NASA Astrophysics Data System (ADS)

    Ambrosio, M. J.; Colavecchia, F. D.; Gasaneo, G.; Mitnik, D. M.; Ancarani, L. U.

    2015-03-01

    The double ionization of helium by high energy electron impact is studied. The corresponding four-body Schrödinger equation is transformed into a set of driven equations containing successive orders in the projectile-target interaction. The first order driven equation is solved with a generalized Sturmian functions approach. The transition amplitude, extracted from the asymptotic limit of the first order solution, is equivalent to the familiar first Born approximation. Fivefold differential cross sections are calculated for (e, 3e) processes within the high incident energy and small momentum transfer regimes. The results are compared with other numerical methods, and with the only absolute experimental data available. Our cross sections agree in shape and magnitude with those of the convergent close coupling method for the (10+10) eV and (4+4) eV emission energies. To date this had not been achieved by any two different numerical schemes when solving the three-body continuum problem for the fast projectile (e, 3e) process. Though agreement with the experimental data, in particular with respect to the magnitude, is not achieved, our findings partly clarify a long standing puzzle.

  18. Determination of optical properties in dental restorative biomaterials using the inverse-adding-doubling method

    NASA Astrophysics Data System (ADS)

    Fernández-Oliveras, Alicia; Rubiño, Manuel; Pérez, María. M.

    2013-11-01

    Light propagation in biological media is characterized by the absorption coefficient, the scattering coefficient, the scattering phase function, the refractive index, and the surface conditions (roughness). By means of the inverse-adding-doubling (IAD) method, transmittance and reflectance measurements lead to the determination of the absorption coefficient and the reduced scattering coefficient. The additional measurement of the phase function performed by goniometry allows the separation of the reduced scattering coefficient into the scattering coefficient and the scattering anisotropy factor. The majority of techniques, such as the one utilized in this work, involve the use of integrating spheres to measure total transmission and reflection. We have employed an integrating sphere setup to measure the total transmittance and reflectance of dental biomaterials used in restorative dentistry. Dental biomaterials are meant to replace dental tissues, such as enamel and dentine, in irreversibly diseased teeth. In previous works we performed goniometric measurements in order to evaluate the scattering anisotropy factor for these kinds of materials. In the present work we have used the IAD method to combine the measurements performed using the integrating sphere setup with the results of the previous goniometric measurements. The aim was to optically characterize the dental biomaterials analyzed, since whole studies to assess the appropriate material properties are required in medical applications. In this context, complete optical characterizations play an important role in achieving the fulfillment of optimal quality and the final success of dental biomaterials used in restorative dentistry.

  19. Evaluation of a method for monoclonal antibody HLA-B27 analysis with the CELL-DYN Sapphire haematology analyser.

    PubMed

    Aarsand, A K; Johannessen, H B; Scott, C S

    2007-12-01

    Analysis of HLA-B27 is usually performed by flow cytometry using commercial single or two colour fluorescence reagents. The CELL-DYN Sapphire (CD-Sapphire) is a high-volume routine haematology analyser that allows cell population analysis by monoclonal antibody fluorochromes analogous to flow cytometry. In this study, in-house flow cytometry analysis (n = 96, HLA-B27, One Lambda) performed on routine patient samples was used as the comparison method for analysis of HLA-B27, One Lambda (n = 40) and HLA-B27/HLA-B7, Immunotech (n = 96) reagents on the CD-Sapphire. The One Lambda results agreed 100% with the comparison method and offered clear population discrimination. The Immunotech combination also had a high level of agreement, but interpretation was more complex because of the wider cross-reactivity of the ABC-m3 antibody with B7 and other HLA-B alleles. When analysing HLA-B27 with antibodies showing nonspecific reactivity, a cut-off staining level yielding high specificity should be chosen, as the primary diagnostic value of HLA-B27 is as a 'rule-out' test for ankylosing spondylitis. The CD-Sapphire incorporates automated sampling and lysis, and medical scientists familiar with the instrument would require little additional technical training to perform the analysis. The reduced preanalytical work and total turnaround time constitute an important step towards automation of HLA-B27 and similar simple high-volume flow cytometry analysis. PMID:17988301

  20. An enzymatic deconjugation method for the analysis of small molecule active drugs on antibody-drug conjugates.

    PubMed

    Li, Yi; Gu, Christine; Gruenhagen, Jason; Yehl, Peter; Chetwyn, Nik P; Medley, Colin D

    2016-01-01

    Antibody-drug conjugates (ADCs) are complex therapeutic agents that use the specific targeting properties of antibodies and the highly potent cytotoxicity of small molecule drugs to selectively eliminate tumor cells while limiting the toxicity to normal healthy tissues. Two critical quality attributes of ADCs are the purity and stability of the active small molecule drug linked to the ADC, but these are difficult to assess once the drug is conjugated to the antibody. In this study, we report a enzyme deconjugation approach to cleave small molecule drugs from ADCs, which allows the drugs to be subsequently characterized by reversed-phase high performance liquid chromatography. The model ADC we used in this study utilizes a valine-citrulline linker that is designed to be sensitive to endoproteases after internalization by tumor cells. We screened several proteases to determine the most effective enzyme. Among the 3 cysteine proteases evaluated, papain had the best efficiency in cleaving the small molecule drug from the model ADC. The deconjugation conditions were further optimized to achieve complete cleavage of the small molecule drug. This papain deconjugation approach demonstrated excellent specificity and precision. The purity and stability of the active drug on an ADC drug product was evaluated and the major degradation products of the active drug were identified. The papain deconjugation method was also applied to several other ADCs, with the results suggesting it could be applied generally to ADCs containing a valine-citrulline linker. Our results indicate that the papain deconjugation method is a powerful tool for characterizing the active small molecule drug conjugated to an ADC, and may be useful in ensuring the product quality, efficacy and the safety of ADCs. PMID:26891281

  1. Automated microaneurysm detection method based on double ring filter in retinal fundus images

    NASA Astrophysics Data System (ADS)

    Mizutani, Atsushi; Muramatsu, Chisako; Hatanaka, Yuji; Suemori, Shinsuke; Hara, Takeshi; Fujita, Hiroshi

    2009-02-01

    The presence of microaneurysms in the eye is one of the early signs of diabetic retinopathy, which is one of the leading causes of vision loss. We have been investigating a computerized method for the detection of microaneurysms on retinal fundus images, which were obtained from the Retinopathy Online Challenge (ROC) database. The ROC provides 50 training cases, in which "gold standard" locations of microaneurysms are provided, and 50 test cases without the gold standard locations. In this study, the computerized scheme was developed by using the training cases. Although the results for the test cases are also included, this paper mainly discusses the results for the training cases because the "gold standard" for the test cases is not known. After image preprocessing, candidate regions for microaneurysms were detected using a double-ring filter. Any potential false positives located in the regions corresponding to blood vessels were removed by automatic extraction of blood vessels from the images. Twelve image features were determined, and the candidate lesions were classified into microaneurysms or false positives using the rule-based method and an artificial neural network. The true positive fraction of the proposed method was 0.45 at 27 false positives per image. Forty-two percent of microaneurysms in the 50 training cases were considered invisible by the consensus of two co-investigators. When the method was evaluated for visible microaneurysms, the sensitivity for detecting microaneurysms was 65% at 27 false positives per image. Our computerized detection scheme could be improved for helping ophthalmologists in the early diagnosis of diabetic retinopathy.

  2. A simple in vitro method of evaluating monoclonal antibody binding to tumor tissue

    SciTech Connect

    Rusckowski, M.; Hnatowich, D.J.; Doherty, P.W.; Virzi, F.; Bogden, A.E.

    1984-01-01

    Application of hybridoma technology has resulted in the availability of many monoclonal antibodies (MoAbs) which are potentially useful for in vivo tumor localization in cancer diagnosis and therapy. What is now required is a simple and predictive assay which will select those MoAbs with superior properties for clinical trials. The authors have developed a simple in vitro means of evaluating the relative binding properties of a panel of MoAbs to human tumor tissue. Fresh and frozen surgical explants of human colorectal and breast tumor tissue as 1 mm cubes were incubated in microtiter-plate wells containing media and radio-labelled MoAb. A MoAb specific for carcinoembryonic antigen (anti-CEA) was used as a known colorectal-specific MoAb and an anti-prostatic acid phosphatase MoAb (anti-PAP) was chosen to measure non-specific binding. The MoAbs were coupled with DTPA and labeled with /sup 111/In. Kinetic parameters were measured which define the rate of MoAb uptake and levels of saturation in tissue. In colorectal tumor tissue the anti-CEA:anti-PAP binding ratio was about 10 while in breast tumor tissue this value was obtained by saturating the tumor antigenic sites with excess unlabeled anti-CEA: the levels of binding of specific MoAb was reduced to that of the control MoAb. Equivalent results were obtained with the same tissue which had been stored at -70/sup 0/C for 24 hours. Evaluation of 7 anti-breast and 4 anti-colorectal MoAbs in their respective tumor tissues showed good reproducibility of repeat measurements and 1-15 fold differences in binding between different antibodies in the same tissue. The authors' results suggest that this in vitro assay may help to identify superior MoAb for human trials.

  3. Comparison of Ensemble and Single Molecule Methods for Particle Characterization and Binding Analysis of a PEGylated Single-Domain Antibody.

    PubMed

    Schneeweis, Lumelle A; Obenauer-Kutner, Linda; Kaur, Parminder; Yamniuk, Aaron P; Tamura, James; Jaffe, Neil; O'Mara, Brian W; Lindsay, Stuart; Doyle, Michael; Bryson, James

    2015-12-01

    Domain antibodies (dAbs) are single immunoglobulin domains that form the smallest functional unit of an antibody. This study investigates the behavior of these small proteins when covalently attached to the polyethylene glycol (PEG) moiety that is necessary for extending the half-life of a dAb. The effect of the 40 kDa PEG on hydrodynamic properties, particle behavior, and receptor binding of the dAb has been compared by both ensemble solution and surface methods [light scattering, isothermal titration calorimetry (ITC), surface Plasmon resonance (SPR)] and single-molecule atomic force microscopy (AFM) methods (topography, recognition imaging, and force microscopy). The large PEG dominates the properties of the dAb-PEG conjugate such as a hydrodynamic radius that corresponds to a globular protein over four times its size and a much reduced association rate. We have used AFM single-molecule studies to determine the mechanism of PEG-dependent reductions in the effectiveness of the dAb observed by SPR kinetic studies. Recognition imaging showed that all of the PEGylated dAb molecules are active, suggesting that some may transiently become inactive if PEG sterically blocks binding. This helps explain the disconnect between the SPR, determined kinetically, and the force microscopy and ITC results that demonstrated that PEG does not change the binding energy. PMID:26343417

  4. Development and Evaluation of a Direct Fluorescent Antibody Method for the Diagnosis of Pneumocystis carinii Infections in Experimental Animals

    PubMed Central

    Lim, Sook Kyung; Eveland, Warren C.; Porter, Richard J.

    1973-01-01

    Because no fully satisfactory diagnostic method has been available for use in pneumocystis infection, an attempt was made to apply the fluorescent antibody technique in the identification of Pneumocystis carinii. Hyperimmune sera were prepared in rabbits against P. carinii from human and rat sources. After proper adsorption, these antisera were conjugated with fluorescein isothiocyanate and used as reagents in a direct fluorescent antibody procedure. Each of the two reagents was found to stain trypsin-treated P. carinii organisms from either human or rat sources, indicating the presence of common antigens. Stained organisms were demonstrated in the hypopharyngeal material from rats in which pneumocystis infection had been activated by the administration of corticosteroid. From the results reported here, the procedure outlined is considered sufficiently sensitive and specific to justify tests on pneumocystis infections in man. The findings in a series of specimens from human subjects will be reported separately. The method also provides an extended approach to related research problems. The need for controls of the procedure at all points is emphasized. Images PMID:4202704

  5. Symmetrized complex amplitudes for He double photoionization from the time-dependent close coupling and exterior complex scaling methods

    SciTech Connect

    Horner, D.A.; Colgan, J.; Martin, F.; McCurdy, C.W.; Pindzola, M.S.; Rescigno, T.N.

    2004-06-01

    Symmetrized complex amplitudes for the double photoionization of helium are computed by the time-dependent close-coupling and exterior complex scaling methods, and it is demonstrated that both methods are capable of the direct calculation of these amplitudes. The results are found to be in excellent agreement with each other and in very good agreement with results of other ab initio methods and experiment.

  6. HANFORD DOUBLE SHELL TANK (DST) THERMAL & SEISMIC PROJECT BUCKLING EVALUATION METHODS & RESULTS FOR THE PRIMARY TANKS

    SciTech Connect

    MACKEY, T.C.

    2006-03-17

    This report documents a detailed buckling evaluation of the primary tanks in the Hanford double shell waste tanks. The analysis is part of a comprehensive structural review for the Double-Shell Tank Integrity Project. This work also provides information on tank integrity that specifically responds to concerns raise by the Office of Environment, Safety, and Health (ES&H) Oversight (EH-22) during a review (in April and May 2001) of work being performed on the double-shell tank farms, and the operation of the aging waste facility (AWF) primary tank ventilation system.

  7. A Facile Method to Fabricate Double Gyroid as A Polymer Template for Nanohybrids

    NASA Astrophysics Data System (ADS)

    Wang, Hsiao-Fang; Ho, Rong-Ming

    2015-03-01

    Here, we suggest a facile method to acquire double gyroid (DG) phase from the self-assembly of chiral block copolymers (BCPs*), polystyrene- b-poly(L-lactide) (PS-PLLA). A wide region for the formation of DG can be found in the phase diagram of the BCPs*, suggesting that helical phase (H*) from the self-assembly of BCPs* can serve as a stepping stone for the formation of the DG due to an easy path for order-order transition from two-dimensional to three-dimensional (network) structure. Moreover, the order-order transition from metastable H* to stable DG can be expedited by blending the PS-PLLA with compatible entity. Moreover, PS-PLLA blends are prepared by using styrene oligomer (S) to fine-tune the morphologies of the blends at which the molecular weight ratio of the S and compatible PS block (r) is less than 0.1. Owing to the use of the low-molecular-weight oligomer, the increase of BCP chain mobility in the blends significantly reduces the transformation time for the order-order transition from H* to DG. Consequently, nanoporous gyroid SiO2 can be fabricated using hydrolyzed PS-PLLA blends as a template for sol-gel reaction followed by removal of the PS matrix.

  8. Preparation and in vitro evaluation of ethyl cellulose microspheres containing stavudine by the double emulsion method.

    PubMed

    Sahoo, S K; Mallick, A A; Barik, B B; Senapati, P C

    2007-02-01

    The aim of this study was to formulate and evaluate microspheres of stavudine by water-in-oil-in-oil (w/o/o) double emulsion solvent diffusion method using ethyl cellulose and ethyl cellulose in combination with polyvinyl pyrrolidone. A mixed solvent system consisting of acetonitrile and dichloromethane in an 1: 1 ratio and light liquid paraffin was chosen as primary and secondary oil phase, respectively. Span 80 was used as surfactant for stabilizing the secondary oil phase. The influence of formulation factors like stirring speed, surfactant concentration on particle size and polymer:drug ratio and combination of polymers on drug release characteristics of the microspheres was investigated. The prepared microspheres characterized by micrometric properties, drug loading, Fourier transform infrared spectroscopy, X-ray powder difractometry and scanning electron microscopy. The prepared microspheres were white, free flowing and spherical in shape, stable in nature, with 41-65% of drug entrapment efficiency. The best-fit release kinetics was achieved with Higuchi plot followed by first order and zero order. The release of stavudine was influenced by the drug to polymer ratio, particle size and polymer combination. PMID:17341031

  9. Relative clock estimation method between two LEO satellites with a double-difference solution constraint

    NASA Astrophysics Data System (ADS)

    Liu, Junhong; Gu, Defeng; Ju, Bing; Lai, Yuwang; Yi, Dongyun

    2015-04-01

    A method of estimating the relative clocks between two spaceborne global positioning system (GPS) receivers based on the single-difference (SD) observations is investigated in this paper. Especially, the advantages of introducing a double-difference (DD) solution constraint, including the orbits and ambiguities, are discussed with the simulated data and the real data of Gravity Recovery And Climate Experiment (GRACE) satellites. The theoretical accuracy analysis shows that the accuracy of the relative clocks is improved and the edge effects are eliminated with a DD solution constraint. The simulations indicate a potential accuracy improvement of at least 30% of the relative clocks with the constraint. Furthermore, one month's real data is processed and the overlapping data arcs are used to validate the accuracy of the relative clock solutions. The average overlapping root mean square (RMS) of the relative clock solutions is approximate 99 ps and 31 ps without and with the DD solution constraint, respectively. Moreover, the jumps of the day boundaries are weakened evidently by adding the DD solution constraint. This paper demonstrates that the accuracy and stability of the estimated relative clocks between two low earth orbit (LEO) satellites from SD observations are improved obviously with the DD solution constraint.

  10. Double stapling method of anastomosis after esophagectomy with endoscopic stapler to prevent postoperative stricture.

    PubMed

    Murayama, I; Sato, H; Suzuki, T; Ootsuka, Y; Song, K; Yamagata, M; Fukase, T; Iwai, S

    1998-10-01

    To prevent stricture of an anastomotic site after operation of esophageal cancer, a new surgical technique, the "double-stapling method," was designed and applied clinically to 29 patients. According to the surgical technique, an automatic suture device for endoscopy was inserted from the side of the lesser curvature of the stomach to the esophageal side after performing end-side anastomosis between the esophagus and the stomach tube using a conventional circular anastomotic device to perform anastomosis between the anterior wall of the esophagus and the posterior wall of the stomach tube. As a result, a conventional anastomotic site, which was a plane (two dimensional), was transformed into a three-dimensional configuration. In the postoperative measurement of the anastomotic site using a measurement forceps, the inner diameter of the site was 8.6+/-3.1 mm in the circular group, while it was 17.2+/-4.5 mm in the DS group, showing a significant difference (p < 0.0001). Minor leakage was observed in three patients as a postoperative complication, but no postoperative hemorrhage occurred. PMID:9820722

  11. Radiation dose determines the method for quantification of DNA double strand breaks.

    PubMed

    Bulat, Tanja; Keta, Otilija; Korićanac, Lela; Žakula, Jelena; Petrović, Ivan; Ristić-Fira, Aleksandra; Todorović, Danijela

    2016-03-01

    Ionizing radiation induces DNA double strand breaks (DSBs) that trigger phosphorylation of the histone protein H2AX (γH2AX). Immunofluorescent staining visualizes formation of γH2AX foci, allowing their quantification. This method, as opposed to Western blot assay and Flow cytometry, provides more accurate analysis, by showing exact position and intensity of fluorescent signal in each single cell. In practice there are problems in quantification of γH2AX. This paper is based on two issues: the determination of which technique should be applied concerning the radiation dose, and how to analyze fluorescent microscopy images obtained by different microscopes. HTB140 melanoma cells were exposed to γ-rays, in the dose range from 1 to 16 Gy. Radiation effects on the DNA level were analyzed at different time intervals after irradiation by Western blot analysis and immunofluorescence microscopy. Immunochemically stained cells were visualized with two types of microscopes: AxioVision (Zeiss, Germany) microscope, comprising an ApoTome software, and AxioImagerA1 microscope (Zeiss, Germany). Obtained results show that the level of γH2AX is time and dose dependent. Immunofluorescence microscopy provided better detection of DSBs for lower irradiation doses, while Western blot analysis was more reliable for higher irradiation doses. AxioVision microscope containing ApoTome software was more suitable for the detection of γH2AX foci. PMID:26959322

  12. Lagrange-type modeling of continuous dielectric permittivity variation in double-higher-order volume integral equation method

    NASA Astrophysics Data System (ADS)

    Chobanyan, E.; Ilić, M. M.; Notaroš, B. M.

    2015-05-01

    A novel double-higher-order entire-domain volume integral equation (VIE) technique for efficient analysis of electromagnetic structures with continuously inhomogeneous dielectric materials is presented. The technique takes advantage of large curved hexahedral discretization elements—enabled by double-higher-order modeling (higher-order modeling of both the geometry and the current)—in applications involving highly inhomogeneous dielectric bodies. Lagrange-type modeling of an arbitrary continuous variation of the equivalent complex permittivity of the dielectric throughout each VIE geometrical element is implemented, in place of piecewise homogeneous approximate models of the inhomogeneous structures. The technique combines the features of the previous double-higher-order piecewise homogeneous VIE method and continuously inhomogeneous finite element method (FEM). This appears to be the first implementation and demonstration of a VIE method with double-higher-order discretization elements and conformal modeling of inhomogeneous dielectric materials embedded within elements that are also higher (arbitrary) order (with arbitrary material-representation orders within each curved and large VIE element). The new technique is validated and evaluated by comparisons with a continuously inhomogeneous double-higher-order FEM technique, a piecewise homogeneous version of the double-higher-order VIE technique, and a commercial piecewise homogeneous FEM code. The examples include two real-world applications involving continuously inhomogeneous permittivity profiles: scattering from an egg-shaped melting hailstone and near-field analysis of a Luneburg lens, illuminated by a corrugated horn antenna. The results show that the new technique is more efficient and ensures considerable reductions in the number of unknowns and computational time when compared to the three alternative approaches.

  13. Preparation of single or double-network chitosan/poly(vinyl alcohol) gel films through selective cross-linking method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A selective cross-linking method was developed to create single or double network chitosan/poly(vinyl alcohol) gel films. The cross-linking is based on the hydrogen bonding between PVA and borate and the strong electrostatic interaction between chitosan and tripolyphosphate. The resultant gel films ...

  14. Methods of Smoothing Double-Entry Expectancy Tables Applied to the Prediction of Success in College. Research Report No. 91.

    ERIC Educational Resources Information Center

    Kolen, Michael J.; And Others

    Six methods for smoothing double-entry expectancy tables (tables that relate two predictor variables to probability of attaining a selected level of success on a criterion) were compared using data for entering students at 85 colleges and universities. ACT composite scores and self-reported high school grade averages were used to construct…

  15. Imaging of 3-D seismic velocity structure of Southern Sumatra region using double difference tomographic method

    SciTech Connect

    Lestari, Titik; Nugraha, Andri Dian

    2015-04-24

    Southern Sumatra region has a high level of seismicity due to the influence of the subduction system, Sumatra fault, Mentawai fault and stretching zone activities. The seismic activities of Southern Sumatra region are recorded by Meteorological Climatological and Geophysical Agency (MCGA’s) Seismograph network. In this study, we used earthquake data catalog compiled by MCGA for 3013 events from 10 seismic stations around Southern Sumatra region for time periods of April 2009 – April 2014 in order to invert for the 3-D seismic velocities structure (Vp, Vs, and Vp/Vs ratio). We applied double-difference seismic tomography method (tomoDD) to determine Vp, Vs and Vp/Vs ratio with hypocenter adjustment. For the inversion procedure, we started from the initial 1-D seismic velocity model of AK135 and constant Vp/Vs of 1.73. The synthetic travel time from source to receiver was calculated using ray pseudo-bending technique, while the main tomographic inversion was applied using LSQR method. The resolution model was evaluated using checkerboard test and Derivative Weigh Sum (DWS). Our preliminary results show low Vp and Vs anomalies region along Bukit Barisan which is may be associated with weak zone of Sumatran fault and migration of partial melted material. Low velocity anomalies at 30-50 km depth in the fore arc region may indicated the hydrous material circulation because the slab dehydration. We detected low seismic seismicity in the fore arc region that may be indicated as seismic gap. It is coincides contact zone of high and low velocity anomalies. And two large earthquakes (Jambi and Mentawai) also occurred at the contact of contrast velocity.

  16. Imaging of 3-D seismic velocity structure of Southern Sumatra region using double difference tomographic method

    NASA Astrophysics Data System (ADS)

    Lestari, Titik; Nugraha, Andri Dian

    2015-04-01

    Southern Sumatra region has a high level of seismicity due to the influence of the subduction system, Sumatra fault, Mentawai fault and stretching zone activities. The seismic activities of Southern Sumatra region are recorded by Meteorological Climatological and Geophysical Agency (MCGA's) Seismograph network. In this study, we used earthquake data catalog compiled by MCGA for 3013 events from 10 seismic stations around Southern Sumatra region for time periods of April 2009 - April 2014 in order to invert for the 3-D seismic velocities structure (Vp, Vs, and Vp/Vs ratio). We applied double-difference seismic tomography method (tomoDD) to determine Vp, Vs and Vp/Vs ratio with hypocenter adjustment. For the inversion procedure, we started from the initial 1-D seismic velocity model of AK135 and constant Vp/Vs of 1.73. The synthetic travel time from source to receiver was calculated using ray pseudo-bending technique, while the main tomographic inversion was applied using LSQR method. The resolution model was evaluated using checkerboard test and Derivative Weigh Sum (DWS). Our preliminary results show low Vp and Vs anomalies region along Bukit Barisan which is may be associated with weak zone of Sumatran fault and migration of partial melted material. Low velocity anomalies at 30-50 km depth in the fore arc region may indicated the hydrous material circulation because the slab dehydration. We detected low seismic seismicity in the fore arc region that may be indicated as seismic gap. It is coincides contact zone of high and low velocity anomalies. And two large earthquakes (Jambi and Mentawai) also occurred at the contact of contrast velocity.

  17. A simple nonradioactive method for the determination of the binding affinities of antibodies induced by hapten bioconjugates for drugs of abuse.

    PubMed

    Torres, Oscar B; Antoline, Joshua F G; Li, Fuying; Jalah, Rashmi; Jacobson, Arthur E; Rice, Kenner C; Alving, Carl R; Matyas, Gary R

    2016-02-01

    The accurate analytical measurement of binding affinities of polyclonal antibody in sera to heroin, 6-acetylmorphine (6-AM), and morphine has been a challenging task. A simple nonradioactive method that uses deuterium-labeled drug tracers and equilibrium dialysis (ED) combined with ultra performance liquid chromatography/tandem mass spectrometry (UPLC/MS/MS) to measure the apparent dissociation constant (K d) of antibodies to 6-AM and morphine is described. The method can readily detect antibodies with K d in the low nanomolar range. Since heroin is rapidly degraded in sera, esterase inhibitors were included in the assay, greatly reducing heroin hydrolysis. MS/MS detection directly measured the heroin in the assay after overnight ED, thereby allowing the quantitation of % bound heroin in lieu of K d as an alternative measurement to assess heroin binding to polyclonal antibody sera. This is the first report that utilizes a solution-based assay to quantify heroin-antibody binding without being confounded by the presence of 6-AM and morphine and to measure K d of polyclonal antibody to 6-AM. Hapten surrogates 6-AcMorHap, 6-PrOxyHap, MorHap, DiAmHap, and DiPrOxyHap coupled to tetanus toxoid (TT) were used to generate high affinity antibodies to heroin, 6-AM, and morphine. In comparison to competition ED-UPLC/MS/MS which gave K d values in the nanomolar range, the commonly used competition enzyme-linked immunosorbent assay (ELISA) measured the 50% inhibition concentration (IC50) values in the micromolar range. Despite the differences in K d and IC50 values, similar trends in affinities of hapten antibodies to heroin, 6-AM, and morphine were observed by both methods. Competition ED-UPLC/MS/MS revealed that among the five TT-hapten bioconjugates, TT-6-AcMorHap and TT-6-PrOxyHap induced antibodies that bound heroin, 6-AM, and morphine. In contrast, TT-MorHap induced antibodies that poorly bound heroin, while TT-DiAmHap and TT-DiPrOxyHap induced antibodies either did not

  18. Sensitivity enhancement of 29Si double-quantum dipolar recoupling spectroscopy by Carr-Purcell-Meiboom-Gill acquisition method

    NASA Astrophysics Data System (ADS)

    Goswami, M.; Madhu, P. K.; Dittmer, J.; Nielsen, N. C.; Ganapathy, S.

    2009-08-01

    An enhancement in the detection sensitivity of dipolar recoupled 29Si double-quantum magic-angle spinning experiment is shown with a Carr-Purcell-Meiboom-Gill (CPMG) train of π pulses during the acquisition period. Symmetry-adapted pulse schemes, such as POST-C7 and SR26411, are used for the double-quantum excitation. Application of POST-C7-CPMG method for framework characterisation is demonstrated in the disordered and catalytically important ZSM-5 molecular sieve. Based on the observed double-quantum dipole-dipole correlations and the known T-site Si connectivities, the assignment of all the signals is made for the orthorhombic phase of the as-synthesised (CN form) material.

  19. Characterization of Monoclonal Antibodies Specific for Erwinia carotovora subsp. atroseptica and Comparison of Serological Methods for Its Sensitive Detection on Potato Tubers.

    PubMed

    Gorris, M T; Alarcon, B; Lopez, M M; Cambra, M

    1994-06-01

    Seven monoclonal antibodies (MAbs) to Erwinia carotovora subsp. atroseptica have been produced. One, called 4G4, reacted with high specificity for serogroup I of E. carotovora subsp. atroseptica, the most common serogroup on potato tubers in different serological assays. Eighty-six strains belonging to different E. carotovora subsp. atroseptica serogroups were assayed. Some strains of serogroup XXII also reacted positively. No cross-reactions were observed against other species of plant pathogenic bacteria or 162 saprophytic bacteria from potato tubers. Only one strain of E. chrysanthemi from potato cross-reacted. A comparison of several serological techniques to detect E. carotovora subsp. atroseptica on potato tubers was performed with MAb 4G4 or polyclonal antibodies. The organism was extracted directly from potato peels of artificially inoculated tubers by soaking or selective enrichment under anaerobiosis in a medium with polypectate. MAb 4G4 was able to detect specifically 240 E. carotovora subsp. atroseptica cells per ml by indirect immunofluorescence and immunofluorescence colony staining and after soaking by ELISA-DAS (double-antibody sandwich enzyme-linked immunosorbent assay) after enrichment. The same amount of cells was detected by using immunolectrotransfer with polyclonal antibodies, and E. carotovora subsp. atroseptica and subsp. carotovora were distinguished by the latter technique. ELISA-DAS using MAb 4G4 with an enrichment step also efficiently detected E. carotovora subsp. atroseptica in naturally infected tubers and plants. PMID:16349293

  20. Efficacy and Toxicity Assessment of Different Antibody Based Antiangiogenic Drugs by Computational Docking Method

    PubMed Central

    Mukherjee, Sayan; Chatterjee, Gopa; Ghosh, Moumita; Das, Bishwajit

    2016-01-01

    Bevacizumab and trastuzumab are two antibody based antiangiogenic drugs that are in clinical practice for the treatment of different cancers. Presently applications of these drugs are based on the empirical choice of clinical experts that follow towards population based clinical trials and, hence, their molecular efficacies in terms of quantitative estimates are not being explored. Moreover, different clinical trials with these drugs showed different toxicity symptoms in patients. Here, using molecular docking study, we made an attempt to reveal the molecular rationale regarding their efficacy and off-target toxicity. Though our study reinforces their antiangiogenic potentiality and, among the two, trastuzumab has much higher efficacy; however, this study also reveals that compared to bevacizumab, trastuzumab has higher toxicity effect, specially on the cardiovascular system. This study also reveals the molecular rationale of ocular dysfunction by antiangiogenic drugs. The molecular rationale of toxicity as revealed in this study may help in the judicious choice as well as therapeutic scheduling of these drugs in different cancers. PMID:27047544

  1. New method to quantitate platelets adhered on biomaterials using monoclonal antibodies to human platelet membrane glycoprotein SZ-21

    SciTech Connect

    Xi, T.F.; Zhang, J.C.; Tian, W.H.; Wang, C.R.; Lei, X.H.; Wai, H.Y.; Ruan, C.G. )

    1990-01-01

    This study developed a new technique to quantitate platelets adhered on biomaterials surfaces in vitro, based on a surface phased radioimmunoassay using a monoclonal antibody SZ-21, directed specifically against the membrane glycoprotein complex IIIa of human platelets. In vitro perfusion is performed in system which consists of testing tubes and infusion pump. After 5 minutes perfusion with fresh ACD anticoagulated human whole blood at 2,000s-1 platelets deposition on surface precoated with proteins determined using anti-human platelet antibody (125 I-SZ-21) are 4,173 +/- 932 (Albumin), 59,032 +/- 25,554 (Fibrinogen), and 71,253 +/- 11,484 (Collagen). Meanwhile, platelets adhered on surfaces of four polymers were determined (platelet/mm2): 19,493 +/- 2,050 (Silicone), 48,193 +/- 4,055 (Polytetrafluoroethylene), 50,375 +/- 8,675 (Polyvinyl chloride) and 101,906 +/- 5,916 (Polyethylene). These results were confirmed by SEM. This method is not only applied for evaluating rapidly and reliably blood compatibility of biomaterials in vitro, but will be used at basic study for interaction of blood materials.

  2. A Quick and Parallel Analytical Method Based on Quantum Dots Labeling for ToRCH-Related Antibodies

    NASA Astrophysics Data System (ADS)

    Yang, Hao; Guo, Qing; He, Rong; Li, Ding; Zhang, Xueqing; Bao, Chenchen; Hu, Hengyao; Cui, Daxiang

    2009-12-01

    Quantum dot is a special kind of nanomaterial composed of periodic groups of II-VI, III-V or IV-VI materials. Their high quantum yield, broad absorption with narrow photoluminescence spectra and high resistance to photobleaching, make them become a promising labeling substance in biological analysis. Here, we report a quick and parallel analytical method based on quantum dots for ToRCH-related antibodies including Toxoplasma gondii, Rubella virus, Cytomegalovirus and Herpes simplex virus type 1 (HSV1) and 2 (HSV2). Firstly, we fabricated the microarrays with the five kinds of ToRCH-related antigens and used CdTe quantum dots to label secondary antibody and then analyzed 100 specimens of randomly selected clinical sera from obstetric outpatients. The currently prevalent enzyme-linked immunosorbent assay (ELISA) kits were considered as “golden standard” for comparison. The results show that the quantum dots labeling-based ToRCH microarrays have comparable sensitivity and specificity with ELISA. Besides, the microarrays hold distinct advantages over ELISA test format in detection time, cost, operation and signal stability. Validated by the clinical assay, our quantum dots-based ToRCH microarrays have great potential in the detection of ToRCH-related pathogens.

  3. Grafted-double walled carbon nanotubes as electrochemical platforms for immobilization of antibodies using a metallic-complex chelating polymer: Application to the determination of adiponectin cytokine in serum.

    PubMed

    Ojeda, Irene; Barrejón, Myriam; Arellano, Luis M; González-Cortés, Araceli; Yáñez-Sedeño, Paloma; Langa, Fernando; Pingarrón, José M

    2015-12-15

    An electrochemical immunosensor for adiponectin (APN) using screen printed carbon electrodes (SPCEs) modified with functionalized double-walled carbon nanotubes (DWCNTs) as platforms for immobilization of the specific antibodies is reported. DWCNTs were functionalized by treatment with 4-aminobenzoic acid (HOOC-Phe) in the presence of isoamylnitrite resulting in the formation of 4-carboxyphenyl-DWCNTs. The oriented binding of specific antibodies toward adiponectin was accomplished by using the metallic-complex chelating polymer Mix&Go™. The HOOC-Phe-DWCNTs-modified SPCEs were characterized by cyclic voltammetry and compared with HOOC-Phe-SWCNTs/SPCE. The different variables affecting the performance of the developed immunosensor were optimized. Under the selected conditions, a calibration plot for APN was constructed showing a range of linearity extending between 0.05 and 10.0 μg/mL which is adequate for the determination of the cytokine in real samples. A detection limit of 14.5 ng/mL was achieved. The so prepared immunosensor exhibited a good reproducibility for the APN measurements, excellent storage stability and selectivity, and a much shorter assay time than the available ELISA kits. The usefulness of the immunosensor for the analysis of real samples was demonstrated by analyzing human serum from female or male healthy patients. PMID:26093125

  4. CD138 and CD31 Double-Positive Cells Comprise the Functional Antibody-Secreting Plasma Cell Compartment in Primate Bone Marrow

    PubMed Central

    Martinez-Murillo, Paola; Pramanik, Lotta; Sundling, Christopher; Hultenby, Kjell; Wretenberg, Per; Spångberg, Mats; Karlsson Hedestam, Gunilla B.

    2016-01-01

    Plasma cells (PCs) are defined as terminally differentiated B cells that secrete large amounts of immunoglobulin (Ig). PCs that reside in the bone marrow (BM) are responsible for maintaining long-term antibody (Ab) responses after infection and vaccination, while PCs present in the blood are generally short-lived. In rhesus macaques, a species frequently used for the evaluation of human vaccines, B cells resemble those found in humans. However, a detailed characterization of BM-resident rhesus PC phenotype and function is lacking. Here, we examined Ig secretion of distinct rhesus CD138+ populations by B cell ELISpot analysis to couple phenotype with function. We demonstrate that the CD20low/−CD138+CD31+ BM population was highly enriched for antibody-secreting cells with IgG being the predominant isotype (60%), followed by IgA (33%) and IgM (7%). Transmission electron microscopy analysis confirmed PC enrichment in the CD20low/−CD138+CD31+ population with cells containing nuclei with “spokes of a wheel” chromatin structure and prominent rough endoplasmic reticulum. This panel also stained human BM PCs and allowed a clear distinction between BM PCs and short-lived peripheral PCs, providing an improved strategy to isolate PCs from rhesus BM for further analysis. PMID:27446073

  5. Linear augmented cylindrical wave method for calculating the electronic structure of double-wall carbon nanotubes

    NASA Astrophysics Data System (ADS)

    D'Yachkov, P. N.; Makaev, D. V.

    2006-10-01

    Electronic structure of double-wall carbon nanotubes (DWNTs) consisting of two concentric graphene cylinders with extremely strong covalent bonding of atoms within the individual graphitic sheets, but very weak van der Waals type interaction between them is calculated in the terms of the linear augmented cylindrical wave (LACW) method. A one-electron potential is used and the approximations are made in the sense of muffin-tin (MT) potentials and local density functional theory only. The atoms of DWNT are considered to be enclosed between cylinder-shaped potential barriers. In this approach, the electronic spectrum of the DWNTs is governed by the free movement of electron in the interatomic space of two cylindrical layers, by electron scattering on the MT spheres, and by electron tunneling between the layers. We have calculated the complete band structures and densities of states in the Fermi level region of the purely semiconducting zigzag DWNTs (n,0)@(n',0) ( 10⩽n⩽23 and 19⩽n'⩽32 ) with interlayer distance 3.2Å⩽Δd⩽3.7Å . Analogously data are obtained for metallic armchair (n,n)@(n',n') nanotubes ( n=5 or 4 and n'=10 or 9). According to the LACW calculations, the interwall coupling results in a distinctly stronger perturbation of the band structure of inner tube as compared to that of the outer one. In the case of semiconducting DWNTs, the minimum gap E11 between the singularities of the conduction and valence bands of the shell tubules decreases from 0.15to0.22eV or increases from 0.7to0.15eV , if dividing n' by three leaves a remainder of 1 or 2, respectively. In both cases, the ΔE11 shifts of the gap do not decay, but slightly oscillate as one goes to the tubules with larger diameters d . For inner tubules, the ΔE11 shift depends strongly on the d . For nmod3=2 series with 10⩽n⩽16 , the shifts ΔE11 are positive, the maximum values of ΔE11 being equal to 0.39 and 0.32eV , respectively. As one goes to the inner tubules with larger diameters

  6. Macrocyclic polyaminocarboxylates for stable radiometal antibody conjugates for therapy, SPECT and PET imaging

    DOEpatents

    Mease, R.C.; Mausner, L.F.; Srivastava, S.C.

    1997-06-17

    A simple method for the synthesis of 1,4,7, 10-tetraazacyclododecane N,N{prime}N{double_prime},N{prime}{double_prime}-tetraacetic acid and 1,4,8,11-tetraazacyclotetradecane N,N{prime},N{double_prime},N{prime}{double_prime}-tetraacetic acid involves cyanomethylating 1,4,7,10-tetraazacyclododecane or 1,4,8,11-tetraazacyclotetradecane to form a tetranitrile and hydrolyzing the tetranitrile. These macrocyclic compounds are functionalized through one of the carboxylates and then conjugated to various biological molecules including monoclonal antibodies. The resulting conjugated molecules are labeled with radiometals for SPECT and PET imaging and for radiotherapy. 4 figs.

  7. Development of a Double Glass Mounting Method Using Formaldehyde Alcohol Azocarmine Lactophenol (FAAL) and its Evaluation for Permanent Mounting of Small Nematodes

    PubMed Central

    ZAHABIUN, Farzaneh; SADJJADI, Seyed Mahmoud; ESFANDIARI, Farideh

    2015-01-01

    Background: Permanent slide preparation of nematodes especially small ones is time consuming, difficult and they become scarious margins. Regarding this problem, a modified double glass mounting method was developed and compared with classic method. Methods: A total of 209 nematode samples from human and animal origin were fixed and stained with Formaldehyde Alcohol Azocarmine Lactophenol (FAAL) followed by double glass mounting and classic dehydration method using Canada balsam as their mounting media. The slides were evaluated in different dates and times, more than four years. Different photos were made with different magnification during the evaluation time. Results: The double glass mounting method was stable during this time and comparable with classic method. There were no changes in morphologic structures of nematodes using double glass mounting method with well-defined and clear differentiation between different organs of nematodes in this method. Conclusion: Using this method is cost effective and fast for mounting of small nematodes comparing to classic method. PMID:26811729

  8. A New Method for Extraction of Double-Stranded RNA from Plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The presence of high molecular weight double-stranded RNA (dsRNA) in plants is associated with the presence of RNA viruses. DsRNA is stable and can be extracted easily from the majority of plant species and provides an excellent tool for characterization of novel viruses that are recalcitrant to pur...

  9. Engineering antibody therapeutics.

    PubMed

    Chiu, Mark L; Gilliland, Gary L

    2016-06-01

    The successful introduction of antibody-based protein therapeutics into the arsenal of treatments for patients has within a few decades fostered intense innovation in the production and engineering of antibodies. Reviewed here are the methods currently used to produce antibodies along with how our knowledge of the structural and functional characterization of immunoglobulins has resulted in the engineering of antibodies to produce protein therapeutics with unique properties, both biological and biophysical, that are leading to novel therapeutic approaches. Antibody engineering includes the introduction of the antibody combining site (variable regions) into a host of architectures including bi and multi-specific formats that further impact the therapeutic properties leading to further advantages and successes in patient treatment. PMID:27525816

  10. Double-peak elution profile of a monoclonal antibody in cation exchange chromatography is caused by histidine-protonation-based charge variants.

    PubMed

    Luo, Haibin; Cao, Mingyan; Newell, Kelcy; Afdahl, Christopher; Wang, Jihong; Wang, William K; Li, Yuling

    2015-12-11

    We have systemically investigated unusual elution behaviors of an IgG4 (mAb A) in cation exchange chromatography (CEX). This mAb A exhibited two elution peaks under certain conditions when being purified by several strong CEX columns. When either of the two peaks was isolated and re-injected on the same column, the similar pattern was observed again during elution. The protein distribution between the two peaks could be altered by NaCl concentration in the feed, or NaCl concentration in wash buffer, or elution pH, suggesting two pH-associated strong-and-weak binding configurations. The protein distributions under different pH values showed good correlation with protonated/un-protonated fractions of a histidine residue. These results suggest that the double-peak elution profile associates with histidine-protonation-based charge variants. By conducting pepsin digestion, amino-acid specific chemical modifications, peptide mapping, and measuring the effects of elution residence time, a histidine in the variable fragment (Fab) was identified to be the root cause. Besides double-peak pattern, mAb A can also exhibit peak-shouldering or single elution peak on different CEX resins, reflecting different resins' resolving capability on protonated/un-protonated forms. This work characterizes a novel cause for unusual elution behaviors in CEX and also provides alternative avenues of purification development for mAbs with similar behaviors. PMID:26596869

  11. Monoclonal Antibodies.

    ERIC Educational Resources Information Center

    Killington, R. A.; Powell, K. L.

    1984-01-01

    Monoclonal antibodies have provided an exciting addition to the "armory" of the molecular biologist and immunologist. This article discusses briefly the concept of, techniques available for, production of, and possible uses of monoclonal antibodies. (Author)

  12. Antimitochondrial antibody

    MedlinePlus

    ... antibodies (AMA) are substances ( antibodies ) that form against mitochondria. The mitochondria are an important part of cells. They are ... often, in people with other kinds of liver disease and some autoimmune diseases. Risks Risks for having ...

  13. Target-selective joint polymerase chain reaction: A robust and rapid method for high-throughput production of recombinant monoclonal antibodies from single cells

    PubMed Central

    2011-01-01

    Background During the development of a therapeutic antibody, large numbers of monoclonal antibodies are required to screen for those that are best suited for the desired activity. Although the single cell-based immunoglobulin variable gene cloning technique is a powerful tool, the current methods remain an obstacle to the rapid production of large numbers of recombinant antibodies. Results We have developed a novel overlap extension polymerase chain reaction, the target-selective joint polymerase chain reaction (TS-jPCR), and applied it to the generation of linear immunoglobulin gene expression constructs. TS-jPCR is conducted using a PCR-amplified immunoglobulin variable gene and an immunoglobulin gene-selective cassette (Ig-cassette) that contains all essential elements for antibody expression and overlapping areas of immunoglobulin gene-specific homology. The TS-jPCR technique is simple and specific; the 3'-random nucleotide-tailed immunoglobulin variable gene fragment and the Ig-cassette are assembled into a linear immunoglobulin expression construct, even in the presence of nonspecifically amplified DNA. We also developed a robotic magnetic beads handling instrument for single cell-based cDNA synthesis to amplify immunoglobulin variable genes by rapid amplification of 5' cDNA ends PCR. Using these methods, we were able to produce recombinant monoclonal antibodies from large numbers of single plasma cells within four days. Conclusion Our system reduces the burden of antibody discovery and engineering by rapidly producing large numbers of recombinant monoclonal antibodies in a short period of time. PMID:21774833

  14. Pseudovirion Particles Bearing Native HIV Envelope Trimers Facilitate a Novel Method for Generating Human Neutralizing Monoclonal Antibodies against HIV

    PubMed Central

    Hicar, Mark D.; Chen, Xuemin; Briney, Bryan; Hammonds, Jason; Wang, Jaang-Jiun; Kalams, Spyros; Spearman, Paul W.; Crowe, James E.

    2010-01-01

    Monomeric HIV envelope vaccines fail to elicit broadly neutralizing antibodies or to protect against infection. Neutralizing antibodies against HIV bind to native, functionally active Env trimers on the virion surface. Gag-Env pseudovirions recapitulate the native trimer, and could serve as an effective epitope presentation platform for study of the neutralizing antibody response in HIV-infected individuals. To address if pseudovirions can recapitulate native HIV virion epitope structures, we carefully characterized these particles, concentrating on the antigenic structure of the coreceptor binding site. By blue native gel shift assays, Gag-Env pseudovirions were shown to contain native trimers that were competent for binding to neutralizing monoclonal antibodies. In ELISA, pseudovirions exhibited increased binding of known CD4-induced antibodies following addition of CD4. Using flow cytometric analysis, fluorescently labeled pseudovirions specifically identified a subset of antigen-specific B cells in HIV-infected subjects. Interestingly, the sequence of one of these novel human antibodies, identified during cloning of single HIV-specific B cells and designated 2C6, exhibited homology to mAb 47e, a known anti-CD4-induced coreceptor binding site antibody. The secreted monoclonal antibody 2C6 did not bind monomeric gp120, but specifically bound envelope on pseudovirions. A recombinant form of the antibody 2C6 acted as a CD4-induced epitope-specific antibody in neutralization assays, yet did not bind monomeric gp120. These findings imply specificity against a quaternary epitope presented on the pseudovirion envelope spike. These data demonstrate that Gag-Env pseudovirions recapitulate CD4 and coreceptor binding pocket antigenic structures and can facilitate identification of B cell clones that secrete neutralizing antibodies. PMID:20531016

  15. Method for Increasing Sensitivity of the Distance Protection on a 330 KV Double-Circuit Transmission Line

    NASA Astrophysics Data System (ADS)

    Brinkis, K.; Vanzovich, E.; Drozd, D.; Mutule, A.

    2010-01-01

    Possibilities of increasing the distance protection (DP) sensitivity are considered for a double-circuit transmission line connected to a substation busbar only with one circuit-breaker in each phase. The DP sensitivity can be increased on such a line using mutual connections of the phase wires along its whole length. The minimal (optimal) number of the connections is found by the proposed calculation method.

  16. A direct qPCR method for residual DNA quantification in monoclonal antibody drugs produced in CHO cells.

    PubMed

    Hussain, Musaddeq

    2015-11-10

    Chinese hamster ovary (CHO) cells are the host cell of choice for manufacturing of monoclonal antibody (mAb) drugs in the biopharmaceutical industry. Host cell DNA is an impurity of such manufacturing process and must be controlled and monitored in order to ensure drug purity and safety. A conventional method for quantification of host residual DNA in drug requires extraction of DNA from the mAb drug substance with subsequent quantification of the extracted DNA using real-time PCR (qPCR). Here we report a method where the DNA extraction step is eliminated prior to qPCR. In this method, which we have named 'direct resDNA qPCR', the mAb drug substance is digested with a protease called KAPA in a 96-well PCR plate, the protease in the digest is then denatured at high temperature, qPCR reagents are added to the resultant reaction wells in the plate along with standards and controls in other wells of the same plate, and the plate subjected to qPCR for analysis of residual host DNA in the samples. This direct resDNA qPCR method for CHO is sensitive to 5.0fg of DNA with high precision and accuracy and has a wide linear range of determination. The method has been successfully tested with four mAbs drug, two IgG1 and two IgG4. Both the purified drug substance as well as a number of process intermediate samples, e.g., bioreactor harvest, Protein A column eluate and ion-exchange column eluates were tested. This method simplifies the residual DNA quantification protocol, reduces time of analysis and leads to increased assay sensitivity and development of automated high-throughput methods. PMID:25850374

  17. Production Of Human Antibodies

    NASA Technical Reports Server (NTRS)

    Sammons, David W.; Neil, Garry A.

    1993-01-01

    Process for making human monoclonal antibodies based on combination of techniques. Antibodies made active against specific antigen. Process involves in vivo immunization of human B lymphocyte cells in mice. B cells of interest enriched in vitro before fusion. Method potentially applicable to any antigen. Does not rely on use of Epstein-Barr virus at any step. Human lymphocytes taken from any source.

  18. A Rapid Method to Characterize Mouse IgG Antibodies and Isolate Native Antigen Binding IgG B Cell Hybridomas.

    PubMed

    Liu, Haolin; White, Janice; Crawford, Frances; Jin, Niyun; Ju, Xiangwu; Liu, Kangtai; Jiang, Chengyu; Marrack, Philippa; Zhang, Gongyi; Kappler, John W

    2015-01-01

    B cell hybridomas are an important source of monoclonal antibodies. In this paper, we developed a high-throughput method to characterize mouse IgG antibodies using surface plasmon resonance technology. This assay rapidly determines their sub-isotypes, whether they bind native antigen and their approximate affinities for the antigen using only 50 μl of hybridoma cell culture supernatant. Moreover, we found that mouse hybridomas secreting IgG antibodies also have membrane form IgG expression without Igα. Based on this surface IgG, we used flow cytometry to isolate rare γ2a isotype switched variants from a γ2b antibody secreting hybridoma cell line. Also, we used fluorescent antigen to single cell sort antigen binding hybridoma cells from bulk mixture of fused hybridoma cells instead of the traditional multi-microwell plate screening and limiting dilution sub-cloning thus saving time and labor. The IgG monoclonal antibodies specific for the native antigen identified with these methods are suitable for in vivo therapeutic uses, but also for sandwich ELISA assays, histology, flow cytometry, immune precipitation and x-ray crystallography. PMID:26317987

  19. An immunocytochemical method for localization of estrogen receptors in rat tissues using a dinitrophenyl (DNP)-labeled rat monoclonal primary antibody.

    PubMed

    Gee, J M; Nicholson, R I; Jasani, B; Newman, G R; Amselgruber, W M

    1990-01-01

    We have developed an immunocytochemical method to demonstrate estrogen receptor in hormone-sensitive tissues of the rat using a dinitrophenyl (DNP) hapten-labeled rat antihuman estrogen receptor monoclonal antibody (MAb), H222. Mouse IgM anti-DNP was used secondarily, followed by a DNP/peroxidase conjugate, diaminobenzidine/hydrogen peroxide chromogen, and silver intensification. This method was applied to tissues from intact female rats and showed that estrogen receptor was localized in the nuclei of the stromal and glandular components of the uterine endometrium. Reduced receptor staining was observed in the luminal epithelium, with minimal myometrial staining. Anterior pituitary glands showed heterogeneous immunostaining and ovaries expressed the receptor predominantly in the interstitial cells; fallopian tubes demonstrated substantial epithelial staining. Uteri from chemically castrated rats showed reduced estrogen receptor immunostaining in both stromal and luminal cells, whereas staining was enhanced in the glandular elements. Classical estrogen-unresponsive tissues (heart, lung, and spleen) were unstained. Antibody controls involved pre-blocking antibody recognition sites on the receptor with unlabeled antibodies to estrogen receptor (H222, H226, and D547), as well as use of an inappropriate DNP-labeled antibody to metallothionein. These controls illustrated the specific nature of the DNP-H222 binding. PMID:1688451

  20. Species- and infective stage-specific monoclonal antibodies to Leishmania major produced by an in vitro immunization method.

    PubMed

    Wu, S J; Rowton, E D; Ma, M; Andre, R G

    1990-12-01

    Monoclonal antibodies specific to the infective-stage promastigotes of Leishmania major are needed for developing rapid diagnostic assays of infected sand flies. An in vitro immunization protocol was applied for the production of monoclonal antibodies using small amounts of L. major. Infective-stage promastigotes were isolated from sand flies (Phlebotomus papatasi) 7-10 days after infection and used as antigen for immunization. Two weeks after a primary immunization, murine splenocytes were removed and immunized in vitro with antigen in murine EL-4 thymoma cell conditioned medium. Three fusions were performed using X63-Ag.653 myeloma cells as fusion partners and two fusions were performed using FOX-NY cells. Antibodies specific to promastigotes were detected using an indirect enzyme-linked immunosorbent assay (ELISA). Initially 56 monoclonal antibodies were selected, and their species and stage specificity were determined using both an ELISA and an indirect fluorescent antibody assay (IFA). Twelve monoclonal antibodies showed species specificity to L. major when tested against four sympatric species of Leishmania. Four other monoclonal antibodies showed species and infective-stage specificity to L. major promastigotes. When tested in immunoblots, all four species- and stage-specific monoclonal antibodies bound to five protein bands that were unique to the infective-stage promastigotes. PMID:2087235

  1. Development of methods based on double Hough transform or Gabor filtering to discriminate between crop and weed in agronomic images

    NASA Astrophysics Data System (ADS)

    Bossu, Jérémie; Gée, Christelle; Guillemin, Jean-Philippe; Truchetet, Frédéric

    2006-02-01

    This paper presents two spatial methods to discriminate between crop and weeds. The application is related to agronomic image with perspective crop rows. The first method uses a double Hough Transform permitting a detection of crop rows and a classification between crop and weeds. The second method is based on Gabor filtering, a band pass filter. The parameters of this filter are detected from a Fast Fourier Transform of the image. For each method, a weed infestation rate is obtained. The two methods are compared and a discussion concludes about the abilities of these methods to detect the crop rows in agronomic images. Finally, we discuss this method regarding the capability of the spatial approach for classifying weeds from crop.

  2. Comparing two methods of plastination and glycerin preservation to study skeletal system after Alizarin red-Alcian blue double staining

    PubMed Central

    Mohsen, Setayesh M.; Esfandiari, Ebrahim; Rabiei, Abbas A.; Hanaei, Mahsa S.; Rashidi, Bahman

    2013-01-01

    Background: Plastination is a new method of preserving tissue samples for a long time. This study aimed to compare the new plastination technique with the conventional preservative method in glycerin for fetus skeleton tissues and young rats dyed by Alizarin red- Alcian blue double staining. Materials and Methods: In this study, 4 groups of 1-day, 3-day, 12-day and mature rats were selected and, after being anesthetized and slaughtered, their skin was completely removed. In Alizarin red- Alcian blue double staining method, first the samples were fixed in 95% ethanol and then their cartilages were dyed by 0.225% Alcian blue solution; after that, they were cleared in 1% KOH. Then, the bones were dyed in 0.003% Alizarin red solution and finally the tissue was decolorized in 95% ethanol. In each group, half of the samples were preserved by the conventional method in a glycerin container and the other half were plastinated. Results: In the present study, the samples preserved by plastination technique were dry, odorless, indecomposable and tangible. Quality of coloring had an inverse relationship with rats’ age. Transparency of the plastinated samples had also an inverse relationship with rats’ age. Therefore, skeletal tissue of younger rats had higher quality and transparency in both preservation methods (glycerin and plastination). Conclusion: This study showed that plastination technique was an appropriate method in comparison with glycerin preservation, which conserved skeletal tissue of fetus and young rats colored by Alizarin red- Alcian blue double staining. And the final result was that plastination technique can generate dry, odorless, indecomposable and tangible samples. PMID:23930264

  3. Determination of the Presence of Diphtheria Toxin in the Myocardial Tissue of Rabbits and a Female Subject by Using an Immunofluorescent Antibody Method

    PubMed Central

    Ceyhan, Mehmet; Ozsurekci, Yasemin; Aydin, Merve M.; Akcali, Kamil Can; Talim, Beril; Celik, Melda; Karadag Oncel, Eda; Gurbuz, Venhar; Aycan, Ahmet Emre; Onbasilar, Ilyas; Buzgan, Turan

    2015-01-01

    Background Clinical diagnosis of diphtheria is often difficult, in particular in countries where the disease is rarely observed, such as Turkey. In 2011, after 12 years of no recorded diphtheria cases in Turkey, a 34-year-old woman was diagnosed with diphtheria; she later died of myocarditis. In this study, we aimed to demonstrate the diagnostic potential of an immunofluorescent antibody method to determine the presence of diphtheria toxin (DT) in the myocardial cells of DT-injected rabbits and the female subject. Methods We randomly divided rabbits into two groups: a control group and a DT-injected group. Diphtheria intoxication was simulated in the rabbits by intravenous injection of DT. The myocardium of the rabbits and the female subject were harvested for histopathologic and immunofluorescence examination. A mouse monoclonal anti-DT antibody was used for the immunofluorescent antibody method. Results The presence of DT in the myocardial cells of both the rabbits and the female subject was visualized using the immunofluorescent method. Conclusions Laboratory diagnosis of diphtheria is challenging because of non-toxigenic C. diphtheriae strains and/or the dysfunction of DT. However, visualizing the presence of DT in the myocardial tissue may act as an indicator of biologically active DT. We validated that an immunofluorescent method, which utilizes a monoclonal anti-DT (A-subunit specific) antibody, is a useful diagnostic tool to determine the presence of DT in the myocardium of rabbits and human. PMID:25883712

  4. A method to study the history of a double oxide film defect in liquid aluminum alloys

    NASA Astrophysics Data System (ADS)

    Raiszadeh, R.; Griffiths, W. D.

    2006-12-01

    Entrained double oxide films have been held responsible for reductions in mechanical properties in aluminum casting alloys. However, their behavior in the liquid metal, once formed, has not been studied directly. It has been proposed that the atmosphere entrapped in the double oxide film defect will continue to react with the liquid metal surrounding it, perhaps leading to its elimination as a significant defect. A silicon-nitride rod with a hole in one end was plunged into liquid aluminum to hold a known volume of air in contact with the liquid metal at a constant temperature. The change in the air volume with time was recorded by real-time X-ray radiography to determine the reaction rates of the trapped atmosphere with the liquid aluminum, creating a model for the behavior of an entrained double oxide film defect. The results from this experiment showed that first oxygen, and then nitrogen, was consumed by the aluminum alloy, to form aluminum oxide and aluminum nitride, respectively. The effect of adding different elements to the liquid aluminum and the effect of different hydrogen contents were also studied.

  5. A novel method for extraction and analysis of gunpowder residues on double-side adhesive coated stubs.

    PubMed

    Zeichner, Arie; Eldar, Baruch

    2004-11-01

    A study was conducted to develop an efficient method for extraction and analysis of gunpowder (propellant) residues from double-side adhesive coated stubs, which are used for sampling suspects or their clothing for gunshot (primer) residues (GSR). Conductive and non-conductive double-side adhesives were examined, and the analysis was carried out by gas chromatography/thermal energy analyzer (GC/TEA) and ion mobility spectrometry (IMS). The optimal procedure for the extraction, as was developed in the present study, employs two stages: (1) extraction of the stubs with a mixture of 80% v/v aqueous solution of 0.1% w/v of sodium azide and 20% v/v of ethanol employing sonication at 80 degrees C for 15 min. and (2) residues from the obtained extract were further extracted with methylene chloride. The methylene chloride phase was concentrated by evaporation prior to analysis. Extraction efficiencies of 30-90% for nitroglycerine (NG) and for 2,4-dinitro toluene (2,4-DNT) were found. No significant interferences in the analysis were observed from the adhesives or skin. Interferences were observed in the analysis by the GC/TEA of the samples collected from hair. The method enables analysis of propellant residues on a double-side adhesive coated stub after it was examined for primer residues by scanning electron microscopy/energy dispersive X-ray spectroscopy (SEM/EDX). Thus, the probative value of the evidence may be increased. PMID:15568690

  6. Antisperm antibodies and in vitro fertilization.

    PubMed

    Janssen, H J; Bastiaans, B A; Goverde, H J; Hollanders, H M; Wetzels, A A; Schellekens, L A

    1992-08-01

    The purpose of this study was to investigate the influence of antisperm antibodies in the male, the female, or both partners on the outcome of in vitro fertilization treatment. The results in terms of ongoing pregnancies in the male and female antibody-positive group were the same as in the antibody-negative group. In the double antibody-positive group two of the three patients became pregnant. When high levels of antisperm antibodies were present on the spermatozoa, the fertilization rate was significantly reduced. In the female positive group no clear relationship between the antibody titer and the fertilization percentage could be detected. Abnormal semen quality was responsible for a much lower fertilization rate than the presence of antibodies. The conclusion of this study is that in vitro fertilization provides an equal change of conception in couples with antisperm antibodies in comparison with couples with no antibodies if the other semen parameters are normal. PMID:1472812

  7. The Kernel Energy Method: Construction of 3 & 4 tuple Kernels from a List of Double Kernel Interactions

    PubMed Central

    Huang, Lulu; Massa, Lou

    2010-01-01

    The Kernel Energy Method (KEM) provides a way to calculate the ab-initio energy of very large biological molecules. The results are accurate, and the computational time reduced. However, by use of a list of double kernel interactions a significant additional reduction of computational effort may be achieved, still retaining ab-initio accuracy. A numerical comparison of the indices that name the known double interactions in question, allow one to list higher order interactions having the property of topological continuity within the full molecule of interest. When, that list of interactions is unpacked, as a kernel expansion, which weights the relative importance of each kernel in an expression for the total molecular energy, high accuracy, and a further significant reduction in computational effort results. A KEM molecular energy calculation based upon the HF/STO3G chemical model, is applied to the protein insulin, as an illustration. PMID:21243065

  8. Grid-based methods for diatomic quantum scattering problems III: Double photoionization of molecular hydrogen in prolate spheroidal coordinates

    SciTech Connect

    Tao, Liang; McCurdy, Bill; Rescigno, Tom

    2010-06-10

    Our previously developed finite-element/ discrete variable representation in prolate spheroidal coordinates is extended to two-electron systems with a study of double ionization of H$_2$ with fixed-nuclei. Particular attention is paid to the development of fast and accurate methods for treating the electron-electron interaction. The use of exterior complex scaling in the implementation offers a simple way of enforcing Coulomb boundary conditions for the electronic double continuum. While the angular distributions calculated in this study are found to be completely consistent with our earlier treatments that employed single-center expansions in spherical coordinates, we find that the magnitude of the integrated cross sections are sensitive to small changes in the initial-state wave function. The present formulation offers significant advantages with respect to convergence and efficiency and opens the way to calculations on more complicated diatomic targets.

  9. Development of an immunoaffinity column method using broad-specificity monoclonal antibodies for simultaneous extraction and cleanup of quinolone and sulfonamide antibiotics in animal muscle tissues

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper describes a novel mixed-bed immunoaffinity column (IAC) method. The IAC was produced by coupling anti-fluoroquinolone and anti-sulfonamide broad-specificity monoclonal antibodies (Mabs) to Sepharose 4B for simultaneously isolating 13 fluoroquinolones (FQs) and 6 sulfonamides (SAs) from s...

  10. Studies on the gonococcal IgA1 protease II. Improved methods of enzyme purification and production of monoclonal antibodies to the enzyme.

    PubMed

    Blake, M S; Eastby, C

    1991-11-22

    Two types of extremely active proteases that cleave human IgA1 are produced by pathogenic Neisseria in minute concentrations. To study the antigenicity of these enzymes, a simplified method is described to purify these enzymes from large batch cultures to obtain a sufficient quantity of these IgA1 proteases to study these characteristics. In addition, we describe the production of both rabbit polyclonal and mouse monoclonal antibodies to one of these enzymes. One such monoclonal antibody seemed directed toward the active site of the IgA1 protease and inhibited its enzymatic activity. PMID:1960418

  11. Rhipicephalus (Boophilus) microplus tick in vitro feeding methods for functional (dsRNA) and vaccine candidate (antibody) screening.

    PubMed

    Lew-Tabor, Ala E; Bruyeres, Anthea G; Zhang, Bing; Rodriguez Valle, Manuel

    2014-09-01

    Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) ticks cause economic losses for cattle industries throughout tropical and subtropical regions of the world estimated at $US2.5 billion annually. Lack of access to efficacious long-lasting vaccination regimes and increases in tick acaricide resistance have led to the investigation of targets for the development of novel tick vaccines and treatments. In vitro tick feeding has been used for many tick species to study the effect of new acaricides on the transmission of tick-borne pathogens. Few studies have reported the use of in vitro feeding for functional genomic studies using RNA interference and/or the effect of specific anti-tick antibodies. In particular, in vitro feeding reports for the cattle tick are limited due to its relatively short hypostome. Previously published methods were further modified to broaden optimal tick sizes/weights, feeding sources including bovine and ovine serum, optimisation of commercially available blood anti-coagulant tubes, and IgG concentrations for effective antibody delivery. Ticks are fed overnight and monitored for ∼5-6 weeks to determine egg output and success of larval emergence using a humidified incubator. Lithium-heparin blood tubes provided the most reliable anti-coagulant for bovine blood feeding compared with commercial citrated (CPDA) and EDTA tubes. Although >30mg semi-engorged ticks fed more reliably, ticks as small as 15mg also fed to repletion to lay viable eggs. Ticks which gained less than ∼10mg during in vitro feeding typically did not lay eggs. One mg/ml IgG from Bm86-vaccinated cattle produced a potent anti-tick effect in vitro (83% efficacy) similar to that observed in vivo. Alternatively, feeding of dsRNA targeting Bm86 did not demonstrate anti-tick effects (11% efficacy) compared with the potent effects of ubiquitin dsRNA. This study optimises R. microplus tick in vitro feeding methods which support the development of cattle tick vaccines and

  12. A novel isolation method for hen egg yolk antibody, "IgY".

    PubMed

    Hatta, H; Kim, M; Yamamoto, T

    1990-10-01

    A method for isolation of egg yolk immunoglobulin, IgG, a livetin protein, was investigated. Several natural gums (carrageenan and xanthan gum) were found to be effective for removal of yolk lipoprotein as a precipitate. The effect was pronounced with lambda-carrageenan and the lipid content in the supernatant after removal of the resulting precipitate was less than 0.4% of that of egg yolk. IgY remained in this supernatant, with a yield of 86%, and was isolated by chromatography on a column of DEAE-Sephacel followed by salting-out with sodium sulfate. IgY thus isolated was almost pure (98%) and the yield was 70 to 100 mg per egg. PMID:1368596

  13. Immune Infertility Should Be Positively Diagnosed Using an Accurate Method by Monitoring the Level of Anti-ACTL7a Antibody.

    PubMed

    Fu, Jun; Yao, Rongyan; Luo, Yanyun; Yang, Dantong; Cao, Yang; Qiu, Yi; Song, Wei; Miao, Shiying; Gu, Yiqun; Wang, Linfang

    2016-01-01

    Infertility is currently a major public health problem. Anti-sperm antibodies (ASAs) markedly reduce sperm quality, which can subsequently lead to male and/or female infertility. The accurate detection of ASAs derived from specific spermatozoa is, therefore, clinically useful. We have focused on the spermatozoa-specific expression protein ACTL7a for many years and have developed an enzyme-linked immunosorbent assay (ELISA) to detect the concentration of anti-ACTL7a antibodies in fertile sera (n = 267) and infertile sera (n = 193). Infertile sera were collected from the positive sera of tray agglutination tests (TAT), which is a routine ASA screening methodology. We found that the concentration of anti-ACTL7a antibodies was significantly higher in the infertile sera (than in the fertile sera, P < 0.0001) and much higher in the TAT ≥ 16 infertile sera. The ELISA was much better for male sera detection (AUC = 0.9899). If we set the standard at a strongly positive value (calculated by ROC curve), the positive predictive value of the antibody detection reached 100 percent, with a false positive rate of zero. The developed ELISA method for anti-ACTL7a antibody detection is therefore sensitive, accurate, and easy to perform, making it an excellent potential tool for future clinical use. PMID:26957350

  14. Immune Infertility Should Be Positively Diagnosed Using an Accurate Method by Monitoring the Level of Anti-ACTL7a Antibody

    PubMed Central

    Fu, Jun; Yao, Rongyan; Luo, Yanyun; Yang, Dantong; Cao, Yang; Qiu, Yi; Song, Wei; Miao, Shiying; Gu, Yiqun; Wang, Linfang

    2016-01-01

    Infertility is currently a major public health problem. Anti-sperm antibodies (ASAs) markedly reduce sperm quality, which can subsequently lead to male and/or female infertility. The accurate detection of ASAs derived from specific spermatozoa is, therefore, clinically useful. We have focused on the spermatozoa-specific expression protein ACTL7a for many years and have developed an enzyme-linked immunosorbent assay (ELISA) to detect the concentration of anti-ACTL7a antibodies in fertile sera (n = 267) and infertile sera (n = 193). Infertile sera were collected from the positive sera of tray agglutination tests (TAT), which is a routine ASA screening methodology. We found that the concentration of anti-ACTL7a antibodies was significantly higher in the infertile sera (than in the fertile sera, P < 0.0001) and much higher in the TAT ≥ 16 infertile sera. The ELISA was much better for male sera detection (AUC = 0.9899). If we set the standard at a strongly positive value (calculated by ROC curve), the positive predictive value of the antibody detection reached 100 percent, with a false positive rate of zero. The developed ELISA method for anti-ACTL7a antibody detection is therefore sensitive, accurate, and easy to perform, making it an excellent potential tool for future clinical use. PMID:26957350

  15. Antibody-gold cluster conjugates

    DOEpatents

    Hainfeld, J.F.

    1988-06-28

    Antibody- or antibody fragment-gold cluster conjugates are shown wherein the conjugate size can be about 5.0 nm. Methods and reagents are disclosed in which antibodies or Fab' fragments thereof are covalently bound to a stable cluster of gold atoms. 2 figs.

  16. Novel method for a flexible double-sided microelectrode fabrication process

    NASA Astrophysics Data System (ADS)

    Doerge, Thomas; Kammer, S.; Hanauer, M.; Sossalla, Adam; Steltenkamp, S.

    2009-05-01

    Flexible devices with integrated micro electrodes are widely used for neuronal as well as myogenic stimulation and recording applications. One main intention by using micro electrodes is the ability of placing an appropriate amount of electrodes on the active sites. With an increasing number of single electrodes the selectivity for signal acquirement and analysis is significantly improved. The further advantage of small and elastic structures inside the biological tissue is the perfect fit. This lead to lower traumatisation of the nerve and muscle fibres during and after acute and chronically surgery. Different designed and structured flexible micro electrodes have been developed at the IBMT based on polyimide as substrate material over the last years including cuff, intrafascicular and shaft electrodes. All these systems are generally built up as single sided devices which reduce the possible electrode site half the area. Especially for shaft and intrafascicular applications having double sided electrode arrangement would increase the selectivity enormous. So areas on both sides can be monitored simultaneously. Recent developments of double sided flexible electrode systems lead to promising results especially for varied signal recording. Though these developments revealed some challenges in the field of micromachining including low yield rates. In this work we describe a new technical approach to develop double sided flexible micro electrode systems with a reproducible high yield rate. Prototypes of intrafascicular and intramuscular electrode systems have been developed and investigated by the means of electrochemical characterisation and mechanical behaviour. Additional investigations have been performed with scanning electron microscopy. We also give an outlook to future in vitro and in vivo experiments to investigate the application performance of the developed systems

  17. A Novel Method for Calculation of Strain Energy Release Rate of Asymmetric Double Cantilever Laminated Composite Beams

    NASA Astrophysics Data System (ADS)

    Shokrieh, M. M.; Zeinedini, A.

    2014-06-01

    In this research, a novel data reduction method for calculation of the strain energy release rate ( SERR) of asymmetric double cantilever beams ( ADCB) is presented. For this purpose the elastic beam theory ( EBT) is modified and the new method is called as the modified elastic beam theory ( MEBT). Also, the ADCB specimens are modeled using ABAQUS/Standard software. Then, the initiation of delamination of ADCB specimens is modeled using the virtual crack closure technique ( VCCT). Furthermore, magnitudes of the SERR for different samples are also calculated by an available data reduction method, called modified beam theory ( MBT). Using the hand lay-up method, different laminated composite samples are manufactured by E-glass/epoxy unidirectional plies. In order to measure the SERR, all samples are tested using an experimental setup. The results determined by the new data reduction method ( MEBT) show good agreements with the results of the VCCT and the MBT.

  18. A noniterative asymmetric triple excitation correction for the density-fitted coupled-cluster singles and doubles method: Preliminary applications.

    PubMed

    Bozkaya, Uğur

    2016-04-14

    An efficient implementation of the asymmetric triples correction for the coupled-cluster singles and doubles [ΛCCSD(T)] method [S. A. Kucharski and R. J. Bartlett, J. Chem. Phys. 108, 5243 (1998); T. D. Crawford and J. F. Stanton, Int. J. Quantum Chem. 70, 601 (1998)] with the density-fitting [DF-ΛCCSD(T)] approach is presented. The computational time for the DF-ΛCCSD(T) method is compared with that of ΛCCSD(T). Our results demonstrate that the DF-ΛCCSD(T) method provide substantially lower computational costs than ΛCCSD(T). Further application results show that the ΛCCSD(T) and DF-ΛCCSD(T) methods are very beneficial for the study of single bond breaking problems as well as noncovalent interactions and transition states. We conclude that ΛCCSD(T) and DF-ΛCCSD(T) are very promising for the study of challenging chemical systems, where the coupled-cluster singles and doubles with perturbative triples method fails. PMID:27083709

  19. A noniterative asymmetric triple excitation correction for the density-fitted coupled-cluster singles and doubles method: Preliminary applications

    NASA Astrophysics Data System (ADS)

    Bozkaya, Uǧur

    2016-04-01

    An efficient implementation of the asymmetric triples correction for the coupled-cluster singles and doubles [ΛCCSD(T)] method [S. A. Kucharski and R. J. Bartlett, J. Chem. Phys. 108, 5243 (1998); T. D. Crawford and J. F. Stanton, Int. J. Quantum Chem. 70, 601 (1998)] with the density-fitting [DF-ΛCCSD(T)] approach is presented. The computational time for the DF-ΛCCSD(T) method is compared with that of ΛCCSD(T). Our results demonstrate that the DF-ΛCCSD(T) method provide substantially lower computational costs than ΛCCSD(T). Further application results show that the ΛCCSD(T) and DF-ΛCCSD(T) methods are very beneficial for the study of single bond breaking problems as well as noncovalent interactions and transition states. We conclude that ΛCCSD(T) and DF-ΛCCSD(T) are very promising for the study of challenging chemical systems, where the coupled-cluster singles and doubles with perturbative triples method fails.

  20. Semi-monolithic cavity for external resonant frequency doubling and method of performing the same

    NASA Technical Reports Server (NTRS)

    Hemmati, Hamid (Inventor)

    1999-01-01

    The fabrication of an optical cavity for use in a laser, in a frequency doubling external cavity, or any other type of nonlinear optical device, can be simplified by providing the nonlinear crystal in combination with a surrounding glass having an index of refraction substantially equal to that of the nonlinear crystal. The closed optical path in this cavity is formed in the surrounding glass and through the nonlinear crystal which lies in one of the optical segments of the light path. The light is transmitted through interfaces between the surrounding glass in the nonlinear crystal through interfaces which are formed at the Brewster-angle to minimize or eliminate reflection.

  1. Performances of a method for reconstructing the energy of neutrons detected by a double scattering spectrometer

    SciTech Connect

    Agnello, M.; Botta, E.; Bressani, T.; Calvo, D.; Gianotti, P.; Iazzi, F.; Lamberti, C.; Minetti, B. ); Balocco, E. )

    1992-10-01

    This paper reports on a neutron spectrometer based on the double scattering technique which has been designed and built at the Laboratorio Tecnologico of INFN - Turin (Italy) for Cold Fusion experiments. The operating principle for the reconstruction of the energy can be applied to various fields (neutron emission from sources, fission and fusion) and is described together with the performed tests: a resolution of less than 560 KeV FWHM has been obtained for neutrons of 2.45 MeV, in a typical running configuration.

  2. Ultrastructural localization of specific gonococcal macromolecules with antibody-gold sphere immunological probes.

    PubMed Central

    Robinson, E N; McGee, Z A; Kaplan, J; Hammond, M E; Larson, J K; Buchanan, T M; Schoolnik, G K

    1984-01-01

    In an effort to determine the ultrastructural location of specific macromolecules on the surface of intact microorganisms and in experimentally infected tissues, a new method of rapidly conjugating antibodies to gold spheres via a staphylococcal protein A intermediary has been developed. This new technique provides the excellent density of marking and versatility of sphere size provided by existing gold methods, but decreases preparation time, decreases the chance of bacterial contamination of antibody reagents, and increases specificity of marking. Staphylococcal protein A-coated gold spheres were conjugated with antibodies from rabbits immunized with purified gonococcal pili. The resulting gold-antibody conjugates allowed demonstration of antibody binding to pilus structures of the same gonococcal strain whose pili were used to raise the antibody and demonstration of the lack of antibody recognition of pilus structures on two other gonococcal strains. The failure of gold spheres to attach to isogenic nonpiliated clones of the homologous gonococcus indicated the absence of pilus antigens on the surface of these organisms. The use of a double label--small gold spheres conjugated to anti-pilus antibody and larger gold spheres conjugated to anti-protein I antibody--allowed the simultaneous localization of two gonococcal antigens. Images PMID:6150005

  3. Datasets of a novel bivalent single chain antibody constructed by overlapping oligonucleotide annealing method targeting human CD123.

    PubMed

    Moradi-Kalbolandi, Shima; Habibi-Anbouhi, Mahdi; Golkar, Majid; Behdani, Mahdi; Rezaei, Gashin; Ghazizadeh, Leila; Abolhassani, Mohsen; Shokrgozar, Mohammad Ali

    2016-09-01

    Current therapies for acute myeloid leukemia (AML), are associated with high relapse rates. Hence, development of new therapeutic strategies is crucial to circumvent this problem. Bivalent antibody technology has been used to engineer novel antibody fragments with increased avidity, by assembling two scFv in a single molecule. Here, we present accompanying data from construction and characterization experiments of a biscFv antibody targeting CD123, the most important biomarker of leukemic cancer stem cells which play a key role in relapsed AML after chemotherapy. Data in this article are related to the research paper "Development of a novel engineered antibody targeting human CD123" Moradi-Kalbolandi S. et al. (2016) [1]. PMID:27536714

  4. Toward a Universal Method for Preparing Molecularly Imprinted Polymer Nanoparticles with Antibody-like Affinity for Proteins.

    PubMed

    Xu, Jingjing; Ambrosini, Serena; Tamahkar, Emel; Rossi, Claire; Haupt, Karsten; Tse Sum Bui, Bernadette

    2016-01-11

    We describe a potentially universal, simple and cheap method to prepare water-compatible molecularly imprinted polymer nanoparticles (MIP-NPs) as synthetic antibodies against proteins. The strategy is based on a solid phase synthesis approach where glass beads (GBs) are functionalized with a metal chelate, acting as a general affinity ligand to attract surface-bound histidines present on proteins. This configuration enables an oriented immobilization of the proteins, upon which thermoresponsive MIP-NPs are synthesized. The GBs play the role of both a reactor and a separation column since, after synthesis, the MIP-NPs are released from the support by a simple temperature change, resulting in protein-free polymers. The resulting MIP-NPs are endowed with improved binding site homogeneity, since the binding sites have the same orientation. Moreover, they are stable (no aggregation) in a buffer solution for prolonged storage time and exhibit apparent dissociation constants in the nanomolar range, with little or no cross-reactivity toward other proteins. PMID:26644006

  5. Double photoionization of SO 2 and fragmentation spectroscopy of SO 2++ studied by a photoion-photoion coincidence method

    NASA Astrophysics Data System (ADS)

    Dujardin, Gérald; Leach, Sydney; Dutuit, Odile; Guyon, Paul-Marie; Richard-Viard, Martine

    1984-08-01

    Doubly charged sulphur dioxide cations (SO 2++) are produced by photoionization with synchrotron radiation from ACO in the excitation-energy range 34-54 eV. A new photoion-photoion coincidence (PIPICO) experiment is described in which coincidences between photoion fragments originating from the dissociation of the doubly charged parent cation are counted. This PIPICO method enables us to study the fragmentation of individual electronically excited states of SO 2++ and to determine the corresponding absolute double-photoionization partial cross sections as a function of the excitation energy. A tentative assignment of the three observed α, β and γ SO 2++ states is given. The dissociation processes of the α and β states into the products SO + + O + are found to be non-statistical in nature; the γ state dissociates completely into three atomic fragments S + + O + + O. Three main observed features of the double-photoionization cross-section curves are discussed in the text: appearance potentials, linear threshold laws, and constant double-photoionization cross sections relative to the total ionization cross section at high energies.

  6. Finite Element Method Simulation of Double-Ended Tuning-Fork Quartz Resonator for Application to Vibratory Gyro-Sensor

    NASA Astrophysics Data System (ADS)

    Sato, Kenji; Ono, Atsushi; Tomikawa, Yoshiro

    2003-05-01

    In the present paper, we propose a double-ended tuning-fork quartz resonator for a flatly supported vibratory gyro-sensor in parallel with its rotating plane. The resonator has the advantages of ease of miniaturization and high resistance to external shock, because the height of the proposed resonator is less than that of the conventional vertical-type tuning-fork. In addition, the proposed resonator has two end-support parts. The resonator also has the following features: (1) the vibration energy of the resonator is trapped in the driving part, therefore the resonator is only slightly affected by the support parts and (2) unwanted output signals can be removed by differential connection of the output signals from two symmetric detection electrodes. The resonator was designed using the finite element method (FEM), and its characteristics were also simulated by FEM. The obtained results show that the double-ended tuning-fork quartz resonator is applicable as a vibratory gyro-sensor, and the I/O voltage ratio of the gyro-sensor was found to be proportional to the applied angular velocity. That is, we clarified that the double-ended tuning-fork quartz resonator could be used as a gyro-sensor.

  7. Affinity Purification of Antibodies.

    PubMed

    Hnasko, Robert M; McGarvey, Jeffery A

    2015-01-01

    Antibodies are provided in a variety of formats that include antiserum, hybridoma culture supernatant, or ascites. They can all be used successfully in crude form for the detection of target antigens by immunoassay. However, it is advantageous to use purified antibody in defined quantity to facilitate assay reproducibility, economy, and reduced interference of nonspecific components as well as improved storage, stability, and bio-conjugation. Although not always necessary, the relative simplicity of antibody purification using commercially available protein-A, protein-G, or protein-L resins with basic chromatographic principles warrants purification when antibody source material is available in sufficient quantity. Here, we define three simple methods using immobilized (1) protein-A, (2) protein-G, and (3) protein-L agarose beads to yield highly purified antibody. PMID:26160561

  8. HANFORD DOUBLE SHELL TANK (DST) THERMAL & SEISMIC PROJECT BUCKLING EVALUATION METHODS & RESULTS FOR THE PRIMARY TANKS

    SciTech Connect

    MACKEY TC; JOHNSON KI; DEIBLER JE; PILLI SP; RINKER MW; KARRI NK

    2007-02-14

    This report documents a detailed buckling evaluation of the primary tanks in the Hanford double-shell waste tanks (DSTs), which is part of a comprehensive structural review for the Double-Shell Tank Integrity Project. This work also provides information on tank integrity that specifically responds to concerns raised by the Office of Environment, Safety, and Health (ES&H) Oversight (EH-22) during a review of work performed on the double-shell tank farms and the operation of the aging waste facility (AWF) primary tank ventilation system. The current buckling review focuses on the following tasks: (1) Evaluate the potential for progressive I-bolt failure and the appropriateness of the safety factors that were used for evaluating local and global buckling. The analysis will specifically answer the following questions: (a) Can the EH-22 scenario develop if the vacuum is limited to -6.6-inch water gage (w.g.) by a relief valve? (b) What is the appropriate factor of safety required to protect against buckling if the EH-22 scenario can develop? (c) What is the appropriate factor of safety required to protect against buckling if the EH-22 scenario cannot develop? (2) Develop influence functions to estimate the axial stresses in the primary tanks for all reasonable combinations of tank loads, based on detailed finite element analysis. The analysis must account for the variation in design details and operating conditions between the different DSTs. The analysis must also address the imperfection sensitivity of the primary tank to buckling. (3) Perform a detailed buckling analysis to determine the maximum allowable differential pressure for each of the DST primary tanks at the current specified limits on waste temperature, height, and specific gravity. Based on the I-bolt loads analysis and the small deformations that are predicted at the unfactored limits on vacuum and axial loads, it is very unlikely that the EH-22 scenario (i.e., progressive I-bolt failure leading to global

  9. Fracture Toughness of Thin Plates by the Double-Torsion Test Method

    NASA Technical Reports Server (NTRS)

    Salem, Jonathan A.; Radovic, Miladin; Lara-Curzio, Edgar; Nelson, George

    2006-01-01

    Double torsion testing can produce fracture toughness values without crack length measurement that are comparable to those measured via standardized techniques such as the chevron-notch, surface-crack-in-flexure and precracked beam if the appropriate geometry is employed, and the material does not exhibit increasing crack growth resistance. Results to date indicate that 8 < W/d < 80 and L/W > 2 are required if crack length is not considered in stress intensity calculations. At L/W = 2, the normalized crack length should be 0.35 < a/L < 0.65; whereas for L/W = 3, 0.2 < a/L < 0.75 is acceptable. In addition, the load-points need to roll to reduce friction. For an alumina exhibiting increasing crack growth resistance, values corresponding to the plateau of the R-curve were measured. For very thin plates (W/d > 80) nonlinear effects were encountered.

  10. [Comparison of two types of double-lined simulated landfill leakage detection based on high voltage DC method].

    PubMed

    Yang, Ping; Nai, Chang-Xin; Dong, Lu; Wang, Qi; Wang, Yan-Wen

    2006-01-01

    Two types of double high density polyethylene (HDPE) liners landfill that clay or geogrid was added between the two HDPE liners. The general resistance of the second mode is 15% larger than the general resistance of the first mode in the primary HDPE liner detection, and 20% larger than that of the first one in the secondary HDPE liner detection. High voltage DC method can accomplish the leakage detection and location of these two types of landfill and the error of leakage location is less than 10cm when electrode space is 1m. PMID:16599145

  11. Evaluation of molecular dynamics simulation methods for ionic liquid electric double layers

    NASA Astrophysics Data System (ADS)

    Haskins, Justin B.; Lawson, John W.

    2016-05-01

    We investigate how systematically increasing the accuracy of various molecular dynamics modeling techniques influences the structure and capacitance of ionic liquid electric double layers (EDLs). The techniques probed concern long-range electrostatic interactions, electrode charging (constant charge versus constant potential conditions), and electrolyte polarizability. Our simulations are performed on a quasi-two-dimensional, or slab-like, model capacitor, which is composed of a polarizable ionic liquid electrolyte, [EMIM][BF4], interfaced between two graphite electrodes. To ensure an accurate representation of EDL differential capacitance, we derive new fluctuation formulas that resolve the differential capacitance as a function of electrode charge or electrode potential. The magnitude of differential capacitance shows sensitivity to different long-range electrostatic summation techniques, while the shape of differential capacitance is affected by charging technique and the polarizability of the electrolyte. For long-range summation techniques, errors in magnitude can be mitigated by employing two-dimensional or corrected three dimensional electrostatic summations, which led to electric fields that conform to those of a classical electrostatic parallel plate capacitor. With respect to charging, the changes in shape are a result of ions in the Stern layer (i.e., ions at the electrode surface) having a higher electrostatic affinity to constant potential electrodes than to constant charge electrodes. For electrolyte polarizability, shape changes originate from induced dipoles that soften the interaction of Stern layer ions with the electrode. The softening is traced to ion correlations vertical to the electrode surface that induce dipoles that oppose double layer formation. In general, our analysis indicates an accuracy dependent differential capacitance profile that transitions from the characteristic camel shape with coarser representations to a more diffuse

  12. Chemical etching method assisted double-pulse LIBS for the analysis of silicon crystals

    NASA Astrophysics Data System (ADS)

    Khalil, A. A. I.

    2015-06-01

    Two Nd:YAG lasers working in pulsed modes are combined in the same direction (collinear arrangement) to focus on silicon (Si) crystals in reduced oxygen atmosphere (0.1 mbar) for double-pulse laser-induced breakdown spectroscopy (DP-LIBS) system. Silicon crystals of (100) and (111) orientations were investigated, and Si samples were measured either without prior treatment ("untreated") or after fabrication of nano-pores ("treated"). Nano-pores are produced by metal coating and by chemical etching. DP-LIBS spectra were compared for different Si samples (untreated, treated, (100) and (111) orientations), for double-pulse (DP) (with 266 nm pulse followed by 1064 nm pulse) excitation and for different delay times (times between the excitation laser pulse and the detection ICCD gate); treatment by chemical etching has been studied as well. The intensity of the atomic line Si I at 288.16 nm was enhanced by a factor of about three by using the DP-LIBS signals as compared to the single-pulse (SP) signal which could increase the sensitivity of the LIBS technique. This study proved that an optimized value of the etching time of Si during etching by chemical processes and short delay times are required. Plasma parameters [the electron temperature ( T e) and the electron number density ( N e)] were calculated from measured SP- and DP-LIBS spectra. The most important result of this study is the much higher DP-LIBS intensity observed on Si (100) as compared to Si (111) for measurements under the same experimental conditions. This study could provide important reference data for the design and optimization of DP-LIBS systems involved in plasma-facing components diagnostics.

  13. Evaluation of molecular dynamics simulation methods for ionic liquid electric double layers.

    PubMed

    Haskins, Justin B; Lawson, John W

    2016-05-14

    We investigate how systematically increasing the accuracy of various molecular dynamics modeling techniques influences the structure and capacitance of ionic liquid electric double layers (EDLs). The techniques probed concern long-range electrostatic interactions, electrode charging (constant charge versus constant potential conditions), and electrolyte polarizability. Our simulations are performed on a quasi-two-dimensional, or slab-like, model capacitor, which is composed of a polarizable ionic liquid electrolyte, [EMIM][BF4], interfaced between two graphite electrodes. To ensure an accurate representation of EDL differential capacitance, we derive new fluctuation formulas that resolve the differential capacitance as a function of electrode charge or electrode potential. The magnitude of differential capacitance shows sensitivity to different long-range electrostatic summation techniques, while the shape of differential capacitance is affected by charging technique and the polarizability of the electrolyte. For long-range summation techniques, errors in magnitude can be mitigated by employing two-dimensional or corrected three dimensional electrostatic summations, which led to electric fields that conform to those of a classical electrostatic parallel plate capacitor. With respect to charging, the changes in shape are a result of ions in the Stern layer (i.e., ions at the electrode surface) having a higher electrostatic affinity to constant potential electrodes than to constant charge electrodes. For electrolyte polarizability, shape changes originate from induced dipoles that soften the interaction of Stern layer ions with the electrode. The softening is traced to ion correlations vertical to the electrode surface that induce dipoles that oppose double layer formation. In general, our analysis indicates an accuracy dependent differential capacitance profile that transitions from the characteristic camel shape with coarser representations to a more diffuse

  14. Potential of a spectroscopic measurement method using adding-doubling to retrieve the bulk optical properties of dense microalgal media.

    PubMed

    Bellini, Sarah; Bendoula, Ryad; Latrille, Eric; Roger, Jean-Michel

    2014-01-01

    In the context of algal mass cultivation, current techniques used for the characterization of algal cells require time-consuming sample preparation and a large amount of costly, standard instrumentation. As the physical and chemical properties of the algal cells strongly affect their optical properties, the optical characterization is seen as a promising method to provide an early diagnosis in the context of mass cultivation monitoring. This article explores the potential of a spectroscopic measurement method coupled with the inversion of the radiative transfer theory for the retrieval of the bulk optical properties of dense algal samples. Total transmittance and total reflectance measurements were performed over the 380-1020 nm range on dense algal samples with a double integrating sphere setup. The bulk absorption and scattering coefficients were thus extracted over the 380-1020 nm range by inverting the radiative transfer theory using inverse-adding-doubling computations. The experimental results are presented and discussed; the configuration of the optical setup remains a critical point. The absorption coefficients obtained for the four samples of this study appear not to be more informative about pigment composition than would be classical methods in analytical spectroscopy; however, there is a real added value in measuring the reduced scattering coefficient, as it appears to be strongly correlated to the size distribution of the algal cells. PMID:25198389

  15. Monoclonal antibodies.

    PubMed

    2009-01-01

    The ability to produce and exploit monoclonal antibodies (mAbs) has revolutionized many areas of biological sciences. The unique property of an mAb is that it is a single species of immunoglobulin (IG) molecule. This means that the specificity of the interaction of the paratopes on the IG, with the epitopes on an antigenic target, is the same on every molecule. This property can be used to great benefit in immunoassays to provide tests of defined specificity and sensitivity, which improve the possibilities of standardization. The performance of assays can often be determined relating the actual weight of antibody (hence the number of molecules) to the activity. Often the production of an mAb against a specific epitope is the only way that biological entities can be differentiated. This chapter outlines the areas involving the development of assays based on mAbs. The problems involved address include the physical aspects of mAbs and how they may affect assay design and also the implications of results based on monospecific reagents. Often these are not fully understood, leading to assays that are less than satisfactory, which does not justify the relatively high cost of preparing and screening of mAbs. There are many textbooks and reviews dealing with the preparation of mAbs, the principles involved, and various purification and manipulative methods for the preparation of fragments and conjugation. There has been little general information attempting to summarize the best approaches to assay design using mAbs. Much time can be wasted through bad planning, and this is particularly relevant to mAbs. A proper understanding of some basic principles is essential. It is beyond the scope of this chapter to discuss all aspects, but major areas are highlighted. PMID:19219589

  16. Synthesis of macrocyclic polyaminocarboxylates and their use for preparing stable radiometal antibody immunoconjugates for therapy, SPECT and PET imaging

    DOEpatents

    Mease, R.C.; Mausner, L.F.; Srivastava, S.C.

    1995-06-27

    A simple method for the synthesis of 1,4,7,10-tetraazacyclododecane N,N{prime}N{double_prime},N{prime}{double_prime}-tetraacetic acid and 1,4,8,11-tetraazacyclotetradecane N,N{prime},N{double_prime},N{prime}{double_prime}-tetraacetic acid involves cyanomethylating 1,4,7,10-tetraazacyclododecane or 1,4,8,11-tetraazacyclotetradecane to form a tetranitrile and hydrolyzing the tetranitrile. These macrocyclic compounds are functionalized through one of the carboxylates and then conjugated to various biological molecules including monoclonal antibodies. The resulting conjugated molecules are labeled with radiometals for SPECT and PET imaging and for radiotherapy. 4 figs.

  17. In vitro rabies vaccine potency appraisal by ELISA: advantages of the immunocapture method with a neutralizing anti-glycoprotein monoclonal antibody.

    PubMed

    Perrin, P; Morgeaux, S; Sureau, P

    1990-10-01

    The replacement of the in vivo potency test (NIH test) for rabies vaccine evaluation by in vitro methods is at present discussed in many reports and also by WHO expert working groups. For this purpose, in vitro glycoprotein titration has been proposed. Among the different glycoprotein assays, we have studied two ELISA methods (immunocapture and direct plate coating with the antigen to be tested) using neutralizing mono- and polyclonal antibodies. In our view, the immunocapture method based on the use of a neutralizing monoclonal anti-glycoprotein antibody seems to be a convenient tool for the determination of the in vitro potency of rabies vaccine and of the products corresponding to the different steps of their production process. PMID:2285504

  18. Known plaintext attack on double random phase encoding using fingerprint as key and a method for avoiding the attack.

    PubMed

    Tashima, Hideaki; Takeda, Masafumi; Suzuki, Hiroyuki; Obi, Takashi; Yamaguchi, Masahiro; Ohyama, Nagaaki

    2010-06-21

    We have shown that the application of double random phase encoding (DRPE) to biometrics enables the use of biometrics as cipher keys for binary data encryption. However, DRPE is reported to be vulnerable to known-plaintext attacks (KPAs) using a phase recovery algorithm. In this study, we investigated the vulnerability of DRPE using fingerprints as cipher keys to the KPAs. By means of computational experiments, we estimated the encryption key and restored the fingerprint image using the estimated key. Further, we propose a method for avoiding the KPA on the DRPE that employs the phase retrieval algorithm. The proposed method makes the amplitude component of the encrypted image constant in order to prevent the amplitude component of the encrypted image from being used as a clue for phase retrieval. Computational experiments showed that the proposed method not only avoids revealing the cipher key and the fingerprint but also serves as a sufficiently accurate verification system. PMID:20588510

  19. [Effects of different tillage methods on photosynthetic characteristics, dry matter production and economic benefit of double cropping soybean].

    PubMed

    Tang, Jiang-hua; Su, Li-li; Li, Ya-jie; Xu, Wen-xiu; Peng, Jiang-long

    2016-01-01

    In order to explore suitable mode of high yield cultivation of double cropping soybean after wheat under drip irrigation in northern Xinjiang, field trials were set in 2013-2014 to investigate physiological indices and agronomic traits of double cropping soybean under different tillage methods under drip irrigation. The results showed that leaf area index (LAI), chlorophyll content (SPAD), leaf net photosynthetic rate (Pn), transpiration rate (Tr) and stomatal conductance (g(s)) during the determination period under different tillage methods were in the order of tillage plus film covering (TP)> tillage (T)> rotary tillage (RT) > no-tillage (NT) , and the concentration of intercellular CO₂(Ci) was the opposite. LAI, SPAD, Pn, Tr, and g(s) of TP were higher than that with NT by 55.0%, 9.1%, 41.8%, 37.5% and 56.4%, respectively, and Ci was decreased by 22.1%. TP enhanced the photosynthetic efficiency of soybean and improved the ability of CO₂assimilation, consequently leading to the increase of soybean yield under TP compared to NT. The plant dry matter accumulation of TP treatment was improved greatly, with the pod number and seeds number per plant, 100-seed mass and yield of quadric sowing soybean being increased by 50.3%, 48.1%, 11.8% and 20.8% compared with that under NT, and the differences were significant. Therefore, the plastic film mulching combined with tillage under drip irrigation technology was suitable for double cropping soybean after wheat in northern Xinjiang under this experimental condition. PMID:27228608

  20. Antithyroid microsomal antibody

    MedlinePlus

    Thyroid antimicrosomal antibody; Antimicrosomal antibody; Microsomal antibody; Thyroid peroxidase antibody; TPOAb ... test is done to confirm the cause of thyroid problems, including Hashimoto thyroiditis . The test is also ...

  1. Chlamydia pneumoniae antibody levels before coronary events in the Helsinki Heart Study as measured by different methods.

    PubMed

    Paldanius, Mika; Leinonen, Maija; Virkkunen, Hanna; Tenkanen, Leena; Sävykoski, Tiina; Mänttäri, Matti; Saikku, Pekka

    2006-11-01

    The lack of specific tests for the diagnosis of chronic Chlamydia pneumoniae infection has led to the use of enzyme immunoassay (EIA) instead of the gold standard, that is, microimmunofluorescence (MIF), in the measurement of C. pneumoniae antibodies. We assessed the predictive values of C. pneumoniae antibody levels and seroconversions measured by MIF and EIA for coronary events in the prospective Helsinki Heart Study. Sera from 239 cases with coronary events and 239 controls were available at the baseline and data from 210 cases and 211 controls before and after the event. The agreement between MIF and EIA antibody levels was best in high antibody titers. In conditional logistic regression analysis, only high IgA MIF titers (>/=40) at the baseline predicted future coronary events, and the participants with MIF seroconversion between consecutive sera had a higher (nonsignificant) risk for coronary events than the controls. The difference in the kinetics of EIA and MIF antibodies demonstrated that MIF should remain the gold standard. PMID:16757141

  2. Double-salting out assisted liquid-liquid extraction (SALLE) HPLC method for estimation of temozolomide from biological samples.

    PubMed

    Jain, Darshana; Athawale, Rajani; Bajaj, Amrita; Shrikhande, Shruti

    2014-11-01

    The role of temozolomide (TMZ) in treatment of high grade gliomas, melanomas and other malignancies is being defined by the current clinical developmental trials. Temozolomide belongs to the group of alkylating agents and is prescribed to patients suffering from most aggressive forms of brain tumors. The estimation techniques for temozolomide from the extracted plasma or biological samples includes high-performance liquid chromatography with UV detection (HPLC-UV), micellar electrokinetic capillary chromatography (MKEC) and liquid chromatography coupled to mass spectroscopy (LC-MS). These methods suffer from disadvantages like low resolution, low sensitivity, low recovery or cost involvement. An analytical method possessing capacity to estimate low quantities of TMZ in plasma samples with high extraction efficiency (%) and high resolution with cost effectiveness needs to be developed. Cost effective, robust and low plasma component interfering HPLC method using salting out liquid-liquid extraction (SALLE) technique was developed and validated for estimation of drug from plasma samples. The extraction efficiency (%) with conventional LLE technique with methanol, ethyl acetate, dichloromethane and acetonitrile was found to be 5.99±2.45, 45.39±4.56, 46.04±1.14 and 46.23±3.67 respectively. Extraction efficiency (%) improved with SALLE where sodium chloride was used as an electrolyte and was found to be 6.80±5.56, 52.01±3.13, 62.69±2.11 and 69.20±1.18 with methanol, ethyl acetate, dichloromethane and acetonitrile as organic solvent. Upon utilization of two salts for extraction (double salting liquid-liquid extraction) the extraction efficiency (%) was further improved and was twice of LLE. It was found that double salting liquid-liquid extraction technique yielded extraction efficiency (%) of 11.71±5.66, 55.62±3.44, 77.28±2.89 and 87.75±0.89. Hence a method based on double SALLE was developed for quantification of TMZ demonstrating linearity in the range of

  3. Effect of gamma-irradiation on serum samples on the diagnostic performance of ELISA methods for the detection of trypanosomal antibodies.

    PubMed

    Rebeski, D E; Winger, E M; Gabler, C M; Dwinger, R H; Crowther, J R

    2001-08-01

    The study investigated the effect of gamma-irradiation on bovine serum samples on the ability of enzyme-linked immunosorbent assay (ELISA) methods to detect trypanosomal antibodies. The serum samples were analysed using two standardised indirect ELISA systems. Higher measurement values were observed for most gamma-irradiated antibody positive and negative test samples. Using cut-off points, determined from the analysis of a non-irradiated trypanosomal antibody-negative population, the gamma-irradiated sera data showed that there was an increased risk of misclassifying samples as false positive or cross-reactive due to increased analytical sensitivity and decreased analytical specificity. The intraplate precision and agreement between tested and expected values of measurements were not altered throughout. The impact on the assays' diagnostic performance was estimated by analysing diagnostic sensitivity, diagnostic specificity and related parameters. The data demonstrated that although there was a bias of higher measurement values after gamma-irradiation, this could be compensated after readjustment of the cut-off points to obtain best separation of antibody-positive and -negative samples. Thus, for each assay, no significant difference of the diagnostic proficiency was found before and after gamma-irradiation. The practical implications are discussed of a serum sterilisation procedure using (60)Co gamma-rays for routine sample testing, assay validation and trypanosomosis monitoring and tsetse-fly control and eradication programmes. PMID:11470177

  4. A double-index method to classify Kuroshio intrusion paths in the Luzon Strait

    NASA Astrophysics Data System (ADS)

    Huang, Zhida; Liu, Hailong; Hu, Jianyu; Lin, Pengfei

    2016-06-01

    A double index (DI), which is made up of two sub-indices, is proposed to describe the spatial patterns of the Kuroshio intrusion and mesoscale eddies west to the Luzon Strait, based on satellite altimeter data. The area-integrated negative and positive geostrophic vorticities are defined as the Kuroshio warm eddy index (KWI) and the Kuroshio cold eddy index (KCI), respectively. Three typical spatial patterns are identified by the DI: the Kuroshio warm eddy path (KWEP), the Kuroshio cold eddy path (KCEP), and the leaking path. The primary features of the DI and three patterns are further investigated and compared with previous indices. The effects of the integrated area and the algorithm of the integration are investigated in detail. In general, the DI can overcome the problem of previously used indices in which the positive and negative geostrophic vorticities cancel each other out. Thus, the proportions of missing and misjudged events are greatly reduced using the DI. The DI, as compared with previously used indices, can better distinguish the paths of the Kuroshio intrusion and can be used for further research.

  5. Monoclonal antibodies to gonadotropin subunits

    SciTech Connect

    Ehrlich, P.H.; Moyle, W.R.; Canfield, R.E.

    1985-01-01

    The production of monoclonal antibodies to peptide hormones, with their unifocal binding sites, can provide tools for understanding hormone structure and function. The paper focuses on techniques that are important for the study of monoclonal antibodies to chorionic gonadotropin (hCG), including hybridoma production, methods of screening for desired clones, properties of the monoclonal antibodies, effect of antibodies on hormone-receptor interaction, inhibition of binding of radiolabeled hCG, inhibition of hCG induced steroidogenesis, determination of relative orientation of epitopes, and synergistic actions of monoclonal antibodies to hCG.

  6. Label-free and real-time detection of antigen-antibody interactions by Oblique-incidence Reflectivity Difference (OIRD) method

    NASA Astrophysics Data System (ADS)

    He, LiPing; Sun, Yue; Dai, Jun; Wang, JingYi; Lü, HuiBin; Wang, ShuFang; Jin, KuiJuan; Zhou, YueLiang; Yang, GuoZhen

    2012-09-01

    We label-free and real-time detected three interaction processes of antigen-antibodies, Human Immunoglobulin G (IgG), Rabbit IgG, and Mouse IgG as the targets, and Goat Anti-human IgG, Goat Anti-rabbit IgG, and Goat Anti-mouse IgG as the probe, by the Oblique-incidence Reflectivity Difference (OIRD) method. The interaction dynamic curves of the OIRD signal, corresponding to the interaction processes of antigen-antibodies, are generated. The reaction times from beginning to equilibrium state are about 1800, 900, and 1200 s for Human IgG, Rabbit IgG, and Mouse IgG, respectively. The experimental results demonstrate that the OIRD method not only can distinguish biomolecular interactions, but also can be used in real-time detection of interactions and dynamic processes of biomolecules.

  7. Effects of a Proline Endopeptidase on the Detection and Quantitation of Gluten by Antibody-Based Methods during the Fermentation of a Model Sorghum Beer.

    PubMed

    Panda, Rakhi; Fiedler, Katherine L; Cho, Chung Y; Cheng, Raymond; Stutts, Whitney L; Jackson, Lauren S; Garber, Eric A E

    2015-12-01

    The effectiveness of a proline endopeptidase (PEP) in hydrolyzing gluten and its putative immunopathogenic sequences was examined using antibody-based methods and mass spectrometry (MS). Based on the results of the antibody-based methods, fermentation of wheat gluten containing sorghum beer resulted in a reduction in the detectable gluten concentration. The addition of PEP further reduced the gluten concentration. Only one sandwich ELISA was able to detect the apparent low levels of gluten present in the beers. A competitive ELISA using a pepsin-trypsin hydrolysate calibrant was unreliable because the peptide profiles of the beers were inconsistent with that of the hydrolysate calibrant. Analysis by MS indicated that PEP enhanced the loss of a fragment of an immunopathogenic 33-mer peptide in the beer. However, Western blot results indicated partial resistance of the high molecular weight (HMW) glutenins to the action of PEP, questioning the ability of PEP in digesting all immunopathogenic sequences present in gluten. PMID:26548701

  8. Multiple ion counting ICPMS double spike method for precise U isotopic analysis at ultra-trace levels

    NASA Astrophysics Data System (ADS)

    Snow, Jonathan E.; Friedrich, Jon M.

    2005-04-01

    Of the various methods for the measurement of the isotopic composition of U in solids and solutions, few offer both sensitivity and precision. In recent years, the use of ICPMS technology for this determination has become increasingly prevalent. Here we describe a method for the determination of the 235U/238U ratio in very small quantities (<=350 pg) with an accuracy of better than 3[per mille sign]. We measured several terrestrial standard materials and repeated analyses of the U960 isotopic composition standard. We used a 233U/236U double spike, with multiple ion counting on an unmodified Nu Instruments multicollector ICPMS and a non-standard detector configuration that allows an approximately 20-fold sensitivity gain over the best conventional techniques. This technique shows promise for the detection of isotopic tracers in the environment (for example anthropogenic 238U) at very extreme dilutions, or in cases where the total amount of analyte is necessarily limited.

  9. Incorporation of a QM/MM Buffer Zone in the Variational Double Self-Consistent Field Method

    PubMed Central

    Xie, Wangshen; Song, Lingchun

    2009-01-01

    The explicit polarization (X-Pol) potential is an electronic-structure-based polarization force field, designed for molecular dynamics simulations and modeling of biopolymers. In this approach, molecular polarization and charge transfer effects are explicitly treated by a combined quantum mechanical and molecular mechanical (QM/MM) scheme, and the wave function of the entire system is variationally optimized by a double self-consistent field (DSCF) method. In the present article, we introduce a QM buffer zone for a smooth transition from a QM region to an MM region. Instead of using the Mulliken charge approximation for all QM/MM interactions, the Coulombic interactions between the adjacent fragments are determined directly by electronic structure theory. The present method is designed to accelerate the speed of convergence of the total energy and charge density of the system. PMID:18937511

  10. NMDA receptor antibodies

    PubMed Central

    Ramberger, Melanie; Bsteh, Gabriel; Schanda, Kathrin; Höftberger, Romana; Rostásy, Kevin; Baumann, Matthias; Aboulenein-Djamshidian, Fahmy; Lutterotti, Andreas; Deisenhammer, Florian; Berger, Thomas

    2015-01-01

    Objectives: To analyze the frequency of NMDA receptor (NMDAR) antibodies in patients with various inflammatory demyelinating diseases of the CNS and to determine their clinical correlates. Methods: Retrospective case-control study from 2005 to 2014 with the detection of serum IgG antibodies to NMDAR, aquaporin-4, and myelin oligodendrocyte glycoprotein by recombinant live cell-based immunofluorescence assays. Fifty-one patients with acute disseminated encephalomyelitis, 41 with neuromyelitis optica spectrum disorders, 34 with clinically isolated syndrome, and 89 with multiple sclerosis (MS) were included. Due to a known association of NMDAR antibodies with seizures and behavioral symptoms, patients with those clinical manifestations were preferentially included and are therefore overrepresented in our cohort. Nine patients with NMDAR encephalitis, 94 patients with other neurologic diseases, and 48 healthy individuals were used as controls. Results: NMDAR antibodies were found in all 9 patients with NMDAR encephalitis but in only 1 of 215 (0.5%) patients with inflammatory demyelination and in none of the controls. This patient had relapsing-remitting MS with NMDAR antibodies present at disease onset, with an increase in NMDAR antibody titer with the onset of psychiatric symptoms and cognitive deficits. Conclusion: In demyelinating disorders, NMDAR antibodies are uncommon, even in those with symptoms seen in NMDAR encephalitis. PMID:26309901

  11. Ensemble density functional theory method correctly describes bond dissociation, excited state electron transfer, and double excitations

    SciTech Connect

    Filatov, Michael; Huix-Rotllant, Miquel; Burghardt, Irene

    2015-05-14

    State-averaged (SA) variants of the spin-restricted ensemble-referenced Kohn-Sham (REKS) method, SA-REKS and state-interaction (SI)-SA-REKS, implement ensemble density functional theory for variationally obtaining excitation energies of molecular systems. In this work, the currently existing version of the SA-REKS method, which included only one excited state into the ensemble averaging, is extended by adding more excited states to the averaged energy functional. A general strategy for extension of the REKS-type methods to larger ensembles of ground and excited states is outlined and implemented in extended versions of the SA-REKS and SI-SA-REKS methods. The newly developed methods are tested in the calculation of several excited states of ground-state multi-reference systems, such as dissociating hydrogen molecule, and excited states of donor–acceptor molecular systems. For hydrogen molecule, the new method correctly reproduces the distance dependence of the lowest excited state energies and describes an avoided crossing between the doubly excited and singly excited states. For bithiophene–perylenediimide stacked complex, the SI-SA-REKS method correctly describes crossing between the locally excited state and the charge transfer excited state and yields vertical excitation energies in good agreement with the ab initio wavefunction methods.

  12. Antibody Production with Synthetic Peptides.

    PubMed

    Lee, Bao-Shiang; Huang, Jin-Sheng; Jayathilaka, Lasanthi P; Lee, Jenny; Gupta, Shalini

    2016-01-01

    Peptides (usually 10-20 amino acid residues in length) can be used as effectively as proteins in raising antibodies producing both polyclonal and monoclonal antibodies routinely with titers higher than 20,000. Peptide antigens do not function as immunogens unless they are conjugated to proteins. Production of high quality antipeptide antibodies is dependent upon peptide sequence selection, the success of peptide synthesis, peptide-carrier protein conjugation, the humoral immune response in the host animal, the adjuvant used, the peptide dose administered, the injection method, and the purification of the antibody. Peptide sequence selection is probably the most critical step in the production of antipeptide antibodies. Although the process for designing peptide antigens is not exact, several guidelines and computational B-cell epitope prediction methods can help maximize the likelihood of producing antipeptide antibodies that recognize the protein. Antibodies raised by peptides have become essential tools in life science research. Virtually all phospho-specific antibodies are now produced using phosphopeptides as antigens. Typically, 5-20 mg of peptide is enough for antipeptide antibody production. It takes 3 months to produce a polyclonal antipeptide antibody in rabbits that yields ~100 mL of serum which corresponds to ~8-10 mg of the specific antibody after affinity purification using a peptide column. PMID:27515072

  13. Optimization of a reversed-phase high-performance liquid chromatography/mass spectrometry method for characterizing recombinant antibody heterogeneity and stability.

    PubMed

    Dillon, Thomas M; Bondarenko, Pavel V; Rehder, Douglas S; Pipes, Gary D; Kleemann, Gerd R; Ricci, Margaret Speed

    2006-07-01

    An enhanced analytical RP-HPLC/MS method was developed for monitoring the stability and production of intact and fragmented monoclonal antibodies (MAbs). The use of high column temperatures (70-80 degrees C), organic solvents with high eluotropic strength coefficients (isopropyl and n-propyl alcohols), and Zorbax StableBond columns, were critical for good recovery and resolution of immunoglobulin G1 (IgG1) and IgG2 monoclonal antibodies. Using this method, cleavage products of a degraded IgG1 antibody were clearly separated and identified by in-line electrospray ionization time-of-flight (ESI-TOF) mass spectrometry generating exact masses and unique terminal ladder sequences. The glycosylation profile, including mapping of the terminal galactose and fucose heterogeneity of the N-linked sugars, was determined by mass spectrometry of intact MAbs. In addition, we discovered that several IgG2 MAbs exhibited greater structural heterogeneity compared to IgG1s. Mass spectral characterization data and reduction data suggested that the heterogeneity is disulfide related. This reversed-phase LC/MS method represents a key advancement in monitoring intact MAb production and stability. PMID:16448656

  14. Layered double hydroxide/polyethylene terephthalate nanocomposites. Influence of the intercalated LDH anion and the type of polymerization heating method

    SciTech Connect

    Herrero, M.; Martinez-Gallegos, S.; Labajos, F.M.; Rives, V.

    2011-11-15

    Conventional and microwave heating routes have been used to prepare PET-LDH (polyethylene terephthalate-layered double hydroxide) composites with 1-10 wt% LDH by in situ polymerization. To enhance the compatibility between PET and the LDH, terephthalate or dodecyl sulphate had been previously intercalated in the LDH. PXRD and TEM were used to detect the degree of dispersion of the filler and the type of the polymeric composites obtained, and FTIR spectroscopy confirmed that the polymerization process had taken place. The thermal stability of these composites, as studied by thermogravimetric analysis, was enhanced when the microwave heating method was applied. Dodecyl sulphate was more effective than terephthalate to exfoliate the samples, which only occurred for the terephthalate ones under microwave irradiation. - Graphical abstract: Conventional and microwave heating routes were used to prepare PET-LDH (polyethylene terephthalate-layered double hydroxide) composites with 1-10 wt% LDH by in situ polymerization. To enhance the compatibility between PET and the LDH, terephthalate or dodecyl sulphate was previously intercalated into the LDH. The microwave process improves the dispersion and the thermal stability of nanocomposites due to the interaction of the microwave radiation and the dipolar properties of EG and the homogeneous heating. Highlights: > LDH-PET compatibility is enhanced by preintercalation of organic anions. > Dodecylsulphate performance is much better than that of terephthalate. > Microwave heating improves the thermal stability of the composites. > Microwave heating improves as well the dispersion of the inorganic phase.

  15. Rapid recovery of polycrystalline silicon from kerf loss slurry using double-layer organic solvent sedimentation method

    NASA Astrophysics Data System (ADS)

    Xing, Peng-fei; Guo, Jing; Zhuang, Yan-xin; Li, Feng; Tu, Gan-feng

    2013-10-01

    The rapid development of photovoltaic (PV) industries has led to a shortage of silicon feedstock. However, more than 40% silicon goes into slurry wastes due to the kerf loss in the wafer slicing process. To effectively recycle polycrystalline silicon from the kerf loss slurry, an innovative double-layer organic solvent sedimentation process was presented in the paper. The sedimentation velocities of Si and SiC particles in some organic solvents were investigated. Considering the polarity, viscosity, and density of solvents, the chloroepoxy propane and carbon tetrachloride were selected to separate Si and SiC particles. It is found that Si and SiC particles in the slurry waste can be successfully separated by the double-layer organic solvent sedimentation method, which can greatly reduce the sedimentation time and improve the purity of obtained Si-rich and SiC-rich powders. The obtained Si-rich powders consist of 95.04% Si, and the cast Si ingot has 99.06% Si.

  16. Antithyroglobulin antibody

    MedlinePlus

    ... may be due to: Graves disease Hashimoto thyroiditis Hypothyroidism Systemic lupus erythematosus (SLE) Thyrotoxicosis Type 1 diabetes ... Antibody Chronic thyroiditis (Hashimoto disease) Graves disease Hyperthyroidism Hypothyroidism Systemic lupus erythematosus T3 test Update Date 5/ ...

  17. A PCR-Based Method to Construct Lentiviral Vector Expressing Double Tough Decoy for miRNA Inhibition.

    PubMed

    Qiu, Huiling; Zhong, Jiasheng; Luo, Lan; Liu, Nian; Kang, Kang; Qu, Junle; Peng, Wenda; Gou, Deming

    2015-01-01

    DNA vector-encoded Tough Decoy (TuD) miRNA inhibitor is attracting increased attention due to its high efficiency in miRNA suppression. The current methods used to construct TuD vectors are based on synthesizing long oligonucleotides (~90 mer), which have been costly and problematic because of mutations during synthesis. In this study, we report a PCR-based method for the generation of double Tough Decoy (dTuD) vector in which only two sets of shorter oligonucleotides (< 60 mer) were used. Different approaches were employed to test the inhibitory potency of dTuDs. We demonstrated that dTuD is the most efficient method in miRNA inhibition in vitro and in vivo. Using this method, a mini dTuD library against 88 human miRNAs was constructed and used for a high-throughput screening (HTS) of AP-1 pathway-related miRNAs. Seven miRNAs (miR-18b-5p, -101-3p, -148b-3p, -130b-3p, -186-3p, -187-3p and -1324) were identified as candidates involved in AP-1 pathway regulation. This novel method allows for an accurate and cost-effective generation of dTuD miRNA inhibitor, providing a powerful tool for efficient miRNA suppression in vitro and in vivo. PMID:26624995

  18. Quality Improvement Method using Double Error Correction in Burst Transmission Systems

    NASA Astrophysics Data System (ADS)

    Tsuchiya, Naosuke; Tomiyama, Shigenori; Tanaka, Kimio

    Recently, it has a tendency to reduce an error correction and flow control in order to realize a high speed transmission in a burst transmission systems such as ATM network, IP (Internet Protocol) network, frame relay and so on. Therefore a degradation of network quality, an information loss caused by buffer overflow and decrease of average bit error rate, are occurred, especially for high speed information such as high definition television signals, it is necessary to improve these degradations. This paper proposes one of the typical reconstruction methods of lost information and an improvement of average bit error rate. In order to analyse the degradation phenomena, the Gilbert model is introduced for burst errors and the Fluid flow model for buffer overflow. This method is applied to ATM network which mainly transmit a video signals and it makes clear that proposed method is useful for high speed transmission.

  19. A novel method for evaluating antibody-dependent cell-mediated cytotoxicity by flowcytometry using cryopreserved human peripheral blood mononuclear cells

    PubMed Central

    Yamashita, Makiko; Kitano, Shigehisa; Aikawa, Hiroaki; Kuchiba, Aya; Hayashi, Mitsuhiro; Yamamoto, Noboru; Tamura, Kenji; Hamada, Akinobu

    2016-01-01

    Analyzing the cytotoxic functions of effector cells, such as NK cells against target cancer cells, is thought to be necessary for predicting the clinical efficacy of antibody-dependent cellular cytotoxicity (ADCC) -dependent antibody therapy. The 51Cr release assay has long been the most widely used method for quantification of ADCC activity. However, the reproducibilities of these release assays are not adequate, and they do not allow evaluation of the lysis susceptibilities of distinct cell types within the target cell population. In this study, we established a novel method for evaluating cytotoxicity, which involves the detection and quantification of dead target cells using flowcytometry. CFSE (carboxyfluorescein succinimidyl ester) was used as a dye to specifically stain and thereby label the target cell population, allowing living and dead cells, as well as both target and effector cells, to be quantitatively distinguished. Furthermore, with our new approach, ADCC activity was more reproducibly, sensitively, and specifically detectable, not only in freshly isolated but also in frozen human peripheral blood mononuclear cells (PBMCs), than with the calcein-AM release assay. This assay, validated herein, is expected to become a standard assay for evaluating ADCC activity which will ultimately contribute the clinical development of ADCC dependent-antibody therapies. PMID:26813960

  20. Labeling DNA for Single-Molecule Experiments: Methods of Labeling Internal Specific Sequences on Double-Stranded DNA

    PubMed Central

    Zohar, Hagar; Muller, Susan J.

    2012-01-01

    This review is a practical guide for experimentalists interested in specifically labeling internal sequences on double-stranded (ds) DNA molecules for single-molecule experiments. We describe six labeling approaches demonstrated in a single-molecule context and discuss the merits and drawbacks of each approach with particular attention to the amount of specialized training and reagents required. By evaluating each approach according to criteria relevant to single-molecule experiments, including labeling yield and compatibility with cofactors such as Mg2+, we provide a simple reference for selecting a labeling method for given experimental constraints. Intended for non-specialists seeking accessible solutions to DNA labeling challenges, the approaches outlined emphasize simplicity, robustness, suitability for use by non-biologists, and utility in diverse single-molecule experiments. PMID:21734993

  1. [Study of analgesic efficacy of propacetamol in the postoperative period using a double blind placebo controlled method].

    PubMed

    Nikoda, V V; Maiachkin, R B

    2002-01-01

    The efficiency and safety of postoperative use of propacetamol was estimated in 30 patients by means of double blind placebo controlled method. The first group consisted of 15 patients to whom propacetamol was introduced intravenously in single dose of 2 g along with patient controlled anesthesia with promedol. Placebo in combination with patient control anesthesia were used in 15 patients from the 2nd group. Intravenous introducing of propacetamol in dose of 2 g in 15 minutes provides relief of pain intensity in postoperative period. So it permits to consider propacetamol as basic non-opioid analgesic. In early postoperative period combination of propacetamol and opioid analgesic (promedol) reduces demands in the latter by 44%. PMID:12462772

  2. Labeling DNA for single-molecule experiments: methods of labeling internal specific sequences on double-stranded DNA

    NASA Astrophysics Data System (ADS)

    Zohar, Hagar; Muller, Susan J.

    2011-08-01

    This review is a practical guide for experimentalists interested in specifically labeling internal sequences on double-stranded (ds) DNA molecules for single-molecule experiments. We describe six labeling approaches demonstrated in a single-molecule context and discuss the merits and drawbacks of each approach with particular attention to the amount of specialized training and reagents required. By evaluating each approach according to criteria relevant to single-molecule experiments, including labeling yield and compatibility with cofactors such as Mg2+, we provide a simple reference for selecting a labeling method for given experimental constraints. Intended for non-specialists seeking accessible solutions to DNA labeling challenges, the approaches outlined emphasize simplicity, robustness, suitability for use by non-biologists, and utility in diverse single-molecule experiments.

  3. A hierarchy of local coupled cluster singles and doubles response methods for ionization potentials

    NASA Astrophysics Data System (ADS)

    Wälz, Gero; Usvyat, Denis; Korona, Tatiana; Schütz, Martin

    2016-02-01

    We present a hierarchy of local coupled cluster (CC) linear response (LR) methods to calculate ionization potentials (IPs), i.e., excited states with one electron annihilated relative to a ground state reference. The time-dependent perturbation operator V(t), as well as the operators related to the first-order (with respect to V(t)) amplitudes and multipliers, thus are not number conserving and have half-integer particle rank m. Apart from calculating IPs of neutral molecules, the method offers also the possibility to study ground and excited states of neutral radicals as ionized states of closed-shell anions. It turns out that for comparable accuracy IPs require a higher-order treatment than excitation energies; an IP-CC LR method corresponding to CC2 LR or the algebraic diagrammatic construction scheme through second order performs rather poorly. We therefore systematically extended the order with respect to the fluctuation potential of the IP-CC2 LR Jacobian up to IP-CCSD LR, keeping the excitation space of the first-order (with respect to V(t)) cluster operator restricted to the m = /1 2 ⊕ /3 2 subspace and the accuracy of the zero-order (ground-state) amplitudes at the level of CC2 or MP2. For the more expensive diagrams beyond the IP-CC2 LR Jacobian, we employ local approximations. The implemented methods are capable of treating large molecular systems with hundred atoms or more.

  4. A method for polycrystalline silicon delineation applicable to a double-diffused MOS transistor

    NASA Technical Reports Server (NTRS)

    Halsor, J. L.; Lin, H. C.

    1974-01-01

    Method is simple and eliminates requirement for unreliable special etchants. Structure is graded in resistivity to prevent punch-through and has very narrow channel length to increase frequency response. Contacts are on top to permit planar integrated circuit structure. Polycrystalline shield will prevent creation of inversion layer in isolated region.

  5. Robotic Total Gastrectomy With Intracorporeal Robot-Sewn Anastomosis: A Novel Approach Adopting the Double-Loop Reconstruction Method.

    PubMed

    Parisi, Amilcare; Ricci, Francesco; Trastulli, Stefano; Cirocchi, Roberto; Gemini, Alessandro; Grassi, Veronica; Corsi, Alessia; Renzi, Claudio; De Santis, Francesco; Petrina, Adolfo; Pironi, Daniele; D'Andrea, Vito; Santoro, Alberto; Desiderio, Jacopo

    2015-12-01

    Gastric cancer constitutes a major health problem. Robotic surgery has been progressively developed in this field. Although the feasibility of robotic procedures has been demonstrated, there are unresolved aspects being debated, including the reproducibility of intracorporeal in place of extracorporeal anastomosis.Difficulties of traditional laparoscopy have been described and there are well-known advantages of robotic systems, but few articles in literature describe a full robotic execution of the reconstructive phase while others do not give a thorough explanation how this phase was run.A new reconstructive approach, not yet described in literature, was recently adopted at our Center.Robotic total gastrectomy with D2 lymphadenectomy and a so-called "double-loop" reconstruction method with intracorporeal robot-sewn anastomosis (Parisi's technique) was performed in all reported cases.Preoperative, intraoperative, and postoperative data were collected and a technical note was documented.All tumors were located at the upper third of the stomach, and no conversions or intraoperative complications occurred. Histopathological analysis showed R0 resection obtained in all specimens. Hospital stay was regular in all patients and discharge was recommended starting from the 4th postoperative day. No major postoperative complications or reoperations occurred.Reconstruction of the digestive tract after total gastrectomy is one of the main areas of surgical research in the treatment of gastric cancer and in the field of minimally invasive surgery.The double-loop method is a valid simplification of the traditional technique of construction of the Roux-limb that could increase the feasibility and safety in performing a full hand-sewn intracorporeal reconstruction and it appears to fit the characteristics of the robotic system thus obtaining excellent postoperative clinical outcomes. PMID:26656323

  6. Finite difference method for the arbitrary potential in two dimensions: Application to double/triple quantum dots

    NASA Astrophysics Data System (ADS)

    Ahn, Jai Seok

    2014-01-01

    A finite difference method (FDM) applicable to a two dimensional (2D) quantum dot was developed as a non-conventional approach to the theoretical understandings of quantum devices. This method can be applied to a realistic potential with an arbitrary shape. Using this method, the Hamiltonian in a tri-diagonal matrix could be obtained from any 2D potential, and the Hamiltonian could be diagonalized numerically for the eigenvalues. The legitimacy of this method was first checked by comparing the results with a finite round well with the analytic solutions. Two truncated harmonic wells were examined as a realistic model potential for lateral double quantum dots (DQDs) and for triple quantum dots (TQDs). The successful applications of the 2D FDM were observed with the entanglements in the DQDs. The level-splitting and anticrossing behaviors of the DQDs could be obtained by varying the distance between the dots and by introducing asymmetry in the well-depths. The 2D FDM results for linear/triangular TQDs were compared with the tight binding approximations.

  7. A tracers method for studying double diffusive convection in the liquid layers of planetary interiors

    NASA Astrophysics Data System (ADS)

    Bouffard, M.; Labrosse, M.; Choblet, M.; Fournier, M.; Aubert, M.; Tackley, M.

    2015-10-01

    Convection in the liquid layers of planetary interiors is usually driven by a combination of thermal and compositional sources of buoyancy. The low molecular diffusivity of composition causes troubles in the description of this field on the Eulerian grids typically employed in current codes of geodynamo because numerical diffusion on these grids is potentially larger than the real diffusivity. We developed a Lagrangian description of composition based on a method of tracers. The absence of numerical diffusion inherent to this method allows modeling of thermo-chemical convection with infinite Lewis number. The validation of this new tool on benchmark cases will be presented at EPSC as well as its first applications to the ocean of Ganymede with consistently coupled boundary condi- tions for temperature and composition.

  8. Method of preparing and applying single stranded DNA probes to double stranded target DNAs in situ

    DOEpatents

    Gray, Joe W.; Pinkel, Daniel

    1991-01-01

    A method is provided for producing single stranded non-self-complementary nucleic acid probes, and for treating target DNA for use therewith. Probe is constructed by treating DNA with a restriction enzyme and an exonuclease to form template/primers for a DNA polymerase. The digested strand is resynthesized in the presence of labeled nucleoside triphosphate precursor. Labeled single stranded fragments are separated from the resynthesized fragments to form the probe. Target DNA is treated with the same restriction enzyme used to construct the probe, and is treated with an exonuclease before application of the probe. The method significantly increases the efficiency and specificity of hybridization mixtures by increasing effective probe concentration by eliminating self-hybridization between both probe and target DNAs, and by reducing the amount of target DNA available for mismatched hybridizations.

  9. Method of preparing and applying single stranded DNA probes to double stranded target DNAs in situ

    DOEpatents

    Gray, J.W.; Pinkel, D.

    1991-07-02

    A method is provided for producing single stranded non-self-complementary nucleic acid probes, and for treating target DNA for use therewith. The probe is constructed by treating DNA with a restriction enzyme and an exonuclease to form template/primers for a DNA polymerase. The digested strand is resynthesized in the presence of labeled nucleoside triphosphate precursor. Labeled single stranded fragments are separated from the resynthesized fragments to form the probe. Target DNA is treated with the same restriction enzyme used to construct the probe, and is treated with an exonuclease before application of the probe. The method significantly increases the efficiency and specificity of hybridization mixtures by increasing effective probe concentration by eliminating self-hybridization between both probe and target DNAs, and by reducing the amount of target DNA available for mismatched hybridizations. No Drawings

  10. A New Electrolytic Synthesis Method for Few-Layered MoS2 Nanosheets and Their Robust Biointerfacing with Reduced Antibodies.

    PubMed

    Kukkar, Manil; Tuteja, Satish K; Sharma, Amit L; Kumar, Vinod; Paul, Ashok K; Kim, Ki-Hyun; Sabherwal, Priyanka; Deep, Akash

    2016-07-01

    We report an efficient method for the synthesis of few-layered MoS2 nanosheets and demonstrate their application in the label-free detection of the prostate-specific antigen (PSA) cancer marker. As a novel strategy, the electro-dissolution of molybdenum metal sheets in the presence of Na(+) and S(2-) ions led to the formation of Na(+) intercalated MoS2. Further exfoliation by ultrasonication yielded the desired formation of few-layered MoS2 nanosheets. After comprehensive characterization, the synthesized MoS2 nanosheets were channeled in a field-effect transistor (FET) microdevice. Chemically reduced anti-PSA antibodies were immobilized on the MoS2 channel above the FET microdevice to construct a specific PSA immunosensor. The antibodies were deliberately reduced to expose the hinge-region disulfide bonds. This approach offered a robust and site-directed immunosensing device through biointerfacing of the sulfhydryl groups (-SH) in the reduced antibody with the surface S atoms of MoS2. This device was validated as an effective immunosensor with a low detection limit (10(-5) ng/mL) over a wide linear detection range (10(-5) to 75 ng/mL). PMID:27296984

  11. Accuracy of effective dose estimation in personal dosimetry: a comparison between single-badge and double-badge methods and the MOSFET method.

    PubMed

    Januzis, Natalie; Belley, Matthew D; Nguyen, Giao; Toncheva, Greta; Lowry, Carolyn; Miller, Michael J; Smith, Tony P; Yoshizumi, Terry T

    2014-05-01

    The purpose of this study was three-fold: (1) to measure the transmission properties of various lead shielding materials, (2) to benchmark the accuracy of commercial film badge readings, and (3) to compare the accuracy of effective dose (ED) conversion factors (CF) of the U.S. Nuclear Regulatory Commission methods to the MOSFET method. The transmission properties of lead aprons and the accuracy of film badges were studied using an ion chamber and monitor. ED was determined using an adult male anthropomorphic phantom that was loaded with 20 diagnostic MOSFET detectors and scanned with a whole body CT protocol at 80, 100, and 120 kVp. One commercial film badge was placed at the collar and one at the waist. Individual organ doses and waist badge readings were corrected for lead apron attenuation. ED was computed using ICRP 103 tissue weighting factors, and ED CFs were calculated by taking the ratio of ED and badge reading. The measured single badge CFs were 0.01 (±14.9%), 0.02 (±9.49%), and 0.04 (±15.7%) for 80, 100, and 120 kVp, respectively. Current regulatory ED CF for the single badge method is 0.3; for the double-badge system, they are 0.04 (collar) and 1.5 (under lead apron at the waist). The double-badge system provides a better coefficient for the collar at 0.04; however, exposure readings under the apron are usually negligible to zero. Based on these findings, the authors recommend the use of ED CF of 0.01 for the single badge system from 80 kVp (effective energy 50.4 keV) data. PMID:24670903

  12. Most anti-BrdU antibodies react with 2'-deoxy-5-ethynyluridine -- the method for the effective suppression of this cross-reactivity.

    PubMed

    Liboska, Radek; Ligasová, Anna; Strunin, Dmytro; Rosenberg, Ivan; Koberna, Karel

    2012-01-01

    5-Bromo-2'-deoxyuridine (BrdU) and 2'-deoxy-5-ethynyluridine (EdU) are widely used as markers of replicated DNA. While BrdU is detected using antibodies, the click reaction typically with fluorescent azido-dyes is used for EdU localisation. We have performed an analysis of ten samples of antibodies against BrdU with respect to their reactivity with EdU. Except for one sample all the others evinced reactivity with EdU. A high level of EdU persists in nuclear DNA even after the reaction of EdU with fluorescent azido-dyes if the common concentration of dye is used. Although a ten-time increase of azido-dye concentration resulted in a decrease of the signal provided by anti-BrdU antibodies, it also resulted in a substantial increase of the non-specific signal. We have shown that this unwanted reactivity is effectively suppressed by non-fluorescent azido molecules. In this respect, we have tested two protocols of the simultaneous localisation of incorporated BrdU and EdU. They differ in the mechanism of the revelation of incorporated BrdU for the reaction with antibodies. The first one was based on the use of hydrochloric acid, the second one on the incubation of samples with copper(I) ions. The use of hydrochloric acid resulted in a significant increase of the non-specific signal. In the case of the second method, no such effect was observed. PMID:23272138

  13. Most Anti-BrdU Antibodies React with 2′-Deoxy-5-Ethynyluridine — The Method for the Effective Suppression of This Cross-Reactivity

    PubMed Central

    Strunin, Dmytro; Rosenberg, Ivan; Koberna, Karel

    2012-01-01

    5-Bromo-2′-deoxyuridine (BrdU) and 2′-deoxy-5-ethynyluridine (EdU) are widely used as markers of replicated DNA. While BrdU is detected using antibodies, the click reaction typically with fluorescent azido-dyes is used for EdU localisation. We have performed an analysis of ten samples of antibodies against BrdU with respect to their reactivity with EdU. Except for one sample all the others evinced reactivity with EdU. A high level of EdU persists in nuclear DNA even after the reaction of EdU with fluorescent azido-dyes if the common concentration of dye is used. Although a ten-time increase of azido-dye concentration resulted in a decrease of the signal provided by anti-BrdU antibodies, it also resulted in a substantial increase of the non-specific signal. We have shown that this unwanted reactivity is effectively suppressed by non-fluorescent azido molecules. In this respect, we have tested two protocols of the simultaneous localisation of incorporated BrdU and EdU. They differ in the mechanism of the revelation of incorporated BrdU for the reaction with antibodies. The first one was based on the use of hydrochloric acid, the second one on the incubation of samples with copper(I) ions. The use of hydrochloric acid resulted in a significant increase of the non-specific signal. In the case of the second method, no such effect was observed. PMID:23272138

  14. Rapid double-dye-layer coating for dye-sensitized solar cells using a new method.

    PubMed

    Jung, Cho-long; Han, Chi-Hwan; Moon, Doo Kyung; Jun, Yongseok

    2014-10-01

    Intensive research with the specific aim of developing inexpensive renewable energy sources is currently being undertaken. In dye-sensitized solar cell (DSSC) production, the most time-consuming process is coating the dye on working electrodes: absorption of ruthenium-based dyes [e.g., N719=bis(trtrabutylammonium)-cis-di(thiocyanato)-N,N'-bis(4-carboxylato-4'-carboxylic acid-2,2'-bipyridine) ruthenium(II)] on a photoanode takes a long time. We report a simple dye-coating method using a mixed solvent of ethylene glycol (EG) and glycerol (Gly). According to our experiments, dye-coating time can be reduced to 5 min from several hours. Maximum performance was obtained with an EG/Gly ratio of 1:1. This mixture of solvents gave a performance of 9.1%. Furthermore, the viscous solvent system could control coating depth; positioning dye coatings to a specific depth was rapid and facile. A cell containing two different dyes (N719+black dye) had an efficiency of 9.4%. PMID:25154611

  15. Method and apparatus for controlling crossflow in a double collector main coke oven battery

    SciTech Connect

    Bauer, E.G.

    1986-07-08

    A method is described of controlling the crossflow of gases given off by a coal mass during the production of coke in a coke oven having a coke side collector main and a pusher side collector main comprising the steps of: (a) determining the temperature difference between the temperature in the coke side standpipe and the temperature in the pusher side standpipe, (b) determining the temperature difference between the temperature in the freespace adjacent the coke side of the coke oven and the temperature in the freespace adjacent the pusher side of the coke oven, (c) determining the temperature difference between the temperature of the heating wall of the coke oven adjacent the coke side of the coke oven and the temperature of the heating wall of the coke oven adjacent the pusher side of the coke oven, and (d) opening the coke side standpipe control valve and gooseneck damper and the pusher side standpipe control valve and gooseneck damper, if they are not in the open position, if the temperature difference of step (b) is substantially the same as the temperature difference of step (c) and the temperature difference of step (a) is greater than about 50/sup 0/F in order to control crossflow.

  16. Method and system including a double rotary kiln pyrolysis or gasification of waste material

    DOEpatents

    McIntosh, M.J.; Arzoumanidis, G.G.

    1997-09-02

    A method is described for destructively distilling an organic material in particulate form wherein the particulates are introduced through an inlet into one end of an inner rotating kiln ganged to and coaxial with an outer rotating kiln. The inner and outer kilns define a cylindrical annular space with the inlet being positioned in registry with the axis of rotation of the ganged kilns. During operation, the temperature of the wall of the inner rotary kiln at the inlet is not less than about 500 C to heat the particulate material to a temperature in the range of from about 200 C to about 900 C in a pyrolyzing atmosphere to reduce the particulate material as it moves from the one end toward the other end. The reduced particulates including char are transferred to the annular space between the inner and the outer rotating kilns near the other end of the inner rotating kiln and moved longitudinally in the annular space from near the other end toward the one end in the presence of oxygen to combust the char at an elevated temperature to produce a waste material including ash. Also, heat is provided which is transferred to the inner kiln. The waste material including ash leaves the outer rotating kiln near the one end and the pyrolysis vapor leaves through the particulate material inlet. 5 figs.

  17. Method and system including a double rotary kiln pyrolysis or gasification of waste material

    SciTech Connect

    McIntosh, Michael J.; Arzoumanidis, Gregory G.

    1997-01-01

    A method of destructively distilling an organic material in particulate form wherein the particulates are introduced through an inlet into one end of an inner rotating kiln ganged to and coaxial with an outer rotating kiln. The inner and outer kilns define a cylindrical annular space with the inlet being positioned in registry with the axis of rotation of the ganged kilns. During operation, the temperature of the wall of the inner rotary kiln at the inlet is not less than about 500.degree. C. to heat the particulate material to a temperature in the range of from about 200.degree. C. to about 900.degree. C. in a pyrolyzing atmosphere to reduce the particulate material as it moves from the one end toward the other end. The reduced particulates including char are transferred to the annular space between the inner and the outer rotating kilns near the other end of the inner rotating kiln and moved longitudinally in the annular space from near the other end toward the one end in the presence of oxygen to combust the char at an elevated temperature to produce a waste material including ash. Also, heat is provided which is transferred to the inner kiln. The waste material including ash leaves the outer rotating kiln near the one end and the pyrolysis vapor leaves through the particulate material inlet.

  18. A method and system including a double rotary kiln pyrolysis or gasification of waste material

    SciTech Connect

    McIntosh, M.J.; Arzoumanidis, G.G.

    1995-12-31

    A method is described for destructively distilling an organic material in particulate form wherein the particulates are introduced through an inlet into one end of an inner rotating kiln ganged to and coaxial with an outer rotating kiln. The inner and outer kilns define a cylindrical annular space with the inlet being positioned in registry with the axis of rotation of the ganged kilns. During operation, the temperature of the wall of the inner rotary kiln at the inlet is not less than about 500 C to heat the particulate material to a temperature in the range of from about 200 C to about 900 C in a pyrolyzing atmosphere to reduce the particulate material as it moves from the one end toward the other end. The reduced particulates including char are transferred to the annular space between the inner and the outer rotating kilns near the other end of the inner rotating kiln and moved longitudinally in the annular space from near the other end toward the one end in the presence of oxygen to combust the char at an elevated temperature to produce a waste material including ash. Also, heat is provided which is transferred to the inner kiln. The waste material including ash leaves the outer rotating kiln near the one end and the pyrolysis vapor leaves through the particulate material inlet.

  19. The Double Pedigree: A Method for Studying Culturally and Genetically Inherited Behavior in Tandem

    PubMed Central

    Danchin, Etienne; Pujol, Benoit; Wagner, Richard H.

    2013-01-01

    Transgenerational sources of biological variation have been at the center of evolutionary studies ever since Darwin and Wallace identified natural selection. This is because evolution can only operate on traits whose variation is transmitted, i.e. traits that are heritable. The discovery of genetic inheritance has led to a semantic shift, resulting in the tendency to consider that only genes are inherited across generations. Today, however, concepts of heredity are being broadened again to integrate the accruing evidence of non-genetic inheritance, and many evolutionary biologists are calling for the inclusion of non-genetic inheritance into an inclusive evolutionary synthesis. Here, we focus on social heredity and its role in the inheritance of behavioral traits. We discuss quantitative genetics methods that might allow us to disentangle genetic and non-genetic transmission in natural populations with known pedigrees. We then propose an experimental design based on cross-fostering among animal cultures, environments and families that has the potential to partition inherited phenotypic variation into socially (i.e. culturally) and genetically inherited components. This approach builds towards a new conceptual framework based on the use of an extended version of the animal model of quantitative genetics to integrate genetic and cultural components of behavioral inheritance. PMID:23700404

  20. Bispecific antibodies.

    PubMed

    Kontermann, Roland E; Brinkmann, Ulrich

    2015-07-01

    Bispecific antibodies (bsAbs) combine specificities of two antibodies and simultaneously address different antigens or epitopes. BsAbs with 'two-target' functionality can interfere with multiple surface receptors or ligands associated, for example with cancer, proliferation or inflammatory processes. BsAbs can also place targets into close proximity, either to support protein complex formation on one cell, or to trigger contacts between cells. Examples of 'forced-connection' functionalities are bsAbs that support protein complexation in the clotting cascade, or tumor-targeted immune cell recruiters and/or activators. Following years of research and development (R&D), the first bsAb was approved in 2009. Another bsAb entered the market in December 2014 and several more are in clinical trials. Here, we describe the potentials of bsAbs to become the next wave of antibody-based therapies, focusing on molecules in clinical development. PMID:25728220

  1. Specific antibody for pesticide residue determination produced by antibody-pesticide complex

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A new method for specific antibody production was developed using antibody (Ab)-pesticide complex as a unique immunogen. Parathion (PA) was the targeted pesticide, and rabbit polyclonal antibody (Pab) and mouse monoclonal antibody (Mab) were used as carrier proteins. The Ab-PA complexes were genera...

  2. Implementation of USP antibody standard for system suitability in capillary electrophoresis sodium dodecyl sulfate (CE-SDS) for release and stability methods.

    PubMed

    Esterman, Abbie L; Katiyar, Amit; Krishnamurthy, Girija

    2016-09-01

    Capillary electrophoresis sodium dodecyl sulfate (CE-SDS) is widely used for purity analysis of monoclonal antibody therapeutics for release and stability to demonstrate product consistency and shelf life during the manufacturing and life cycle of the product. CE-SDS method development is focused on exploring the method capability to provide the information about the product purity and product related degradants (fragmentation, aggregation etc.). In order to establish the functionality of the instrumentation, software, and sample preparation; system suitability criteria need to be defined for analytical methods using a well characterized reference standard run under the same protocol and analysis as the test articles. Typically the reference standard is produced using a manufacturing process representative of the clinical material. The qualification, control, and maintenance of in-house reference standards are established through rigorous quality and regulatory guidelines. The U.S. Pharmacopeia (USP) has developed a monoclonal IgG System Suitability Reference Standard to be utilized for assessment of system suitability in CE-SDS methods. In this communication, we evaluate the system suitability acceptance criteria performance of the USP IgG standard using two methods, the recommended USP protocol provided in monograph <129> and a molecule specific Bristol-Myers Squibb (BMS) CE-SDS method. The results from USP IgG standard were compared with two in-house monoclonal antibody reference standards. The data suggest that the USP CE-SDS method may not be suitable for CE-SDS analysis for release and stability of monoclonal antibody therapeutics due to the high level of method induced partial reduction observed for all molecules tested. This high level of fragmentation observed utilizing the USP method will result in reporting lower purity levels, which will impact the overall quality assessment of the molecule. The system suitability criteria recommended by the USP method

  3. Development, evaluation, and in-vivo validation of two non-invasive methods for quantitation of activity and dosimetry of monoclonal antibodies in humans

    SciTech Connect

    Hammond, N.D.; Moldofsky, P.J.; Exten, R.E.; Gatenby, R.A.; Broder, G.J.

    1985-05-01

    The authors have applied both a conjugate view imaging method and a first pass study for quantitation of absolute I-131 activity in lesions and normal tissue of patients with colon carcinoma in order to study biological clearance of the I-131 F(ab)'/sub 2/ fragments of mouse monoclonal antibody and the resultant dosimetry. Both methods require a transmission scan for determining patient attenuation and measurement of patient lesion or organ size in the region of interest. The conjugate view method is analyzed for both SPECT and planar imaging. The percent error of both methods relates to lesion size and absolute activity when compared to actual well-counter assayed samples of malignant and normal tissue obtained from CT-guided needle biopsies or surgical specimens. Dosimetric evaluation was based on determination of activity, clearance from computer-generated time-activity curves and lesion or organ volumes from volumetric CT scan data. The dose to the thyroid gland was calculated for one population receiving Lugol's solution 3 days prior and for the other who received Lugol's at the time of administration. Data showed no significant difference in absorbed thyroid dose. Lastly, the absolute uptake of I-131, lesion to background ratios, and the dosimetry data were compared for three different monoclonal antibody fragments.

  4. A Novel Method for Characterizing Fatigue Delamination Growth Under Mode I Using the Double Cantilever Beam Specimen

    NASA Technical Reports Server (NTRS)

    Carvalho, Nelson; Murri, G.

    2014-01-01

    A novel method is proposed to obtain Mode I delamination growth rate from a Double Cantilever Beam (DCB) specimen. In the proposed method, Unidirectional (UD) DCB specimens are tested in fatigue at different initial maximum energy release rates levels. The growth rate data obtained in the first increments of crack growth at each maximum energy release rate level are used to generate a Paris Law equation, which characterizes delamination growth rate without fiber-bridging, and can also be used to determine a delamination onset curve. The remaining delamination growth rate data from each test are used to determine a modified Paris law, which characterizes the delamination growth rate in a DCB specimen, explicitly accounting for fiber-bridging. The proposed expression captures well the scatter in experimental data obtained using the DCB specimens, suggesting its adequacy. The Paris Law characterizing delamination growth rate without fiber-bridging predicts higher delamination growth rates for the same maximum energy release rate applied, leading to a conservative estimate for delamination growth. This is particularly relevant, since in generic ply interfaces, fiber-bridging is less predominant than in UD DCB specimens. Failing to account for fiber-bridging in UD DCB specimens may underestimate the delamination growth rate, yielding non-conservative predictions.

  5. How the method of synthesis governs the local and global structure of zinc aluminum layered double hydroxides

    SciTech Connect

    Pushparaj, Suraj Shiv Charan; Forano, Claude; Prevot, Vanessa; Lipton, Andrew S.; Rees, Gregory; Hanna, John V.; Nielsen, Ulla Gro

    2015-11-10

    A series of zinc aluminum layered double hydroxides (ZnAl LDHs), [Zn1-xAlx (OH)2Ax,nH2O with A = NO3-, Cl- or CO3] were prepared by the urea and co-precipitation synthesis methods, which allowed for a detailed investigation on how synthesis parameters such as pH, metal ion concentration and post synthesis treatment influence the local and global structure of the LDH product. Information about sample composition, purity, defects and other structural aspects of the LDH products were obtained from powder X-ray diffraction, transmission electron microscopy, micro-Raman, and elemental analysis, as well as solid state 1H, 27Al and 67Zn NMR spectroscopy. Our results show that the urea method results in LDHs, which on the global scale are highly crystalline LDHs, whereas solid state NMR shows the different local environments indicating local disorder most likely linked to the presence of Al-rich phases. However, these Alrich phases are not detected by global range techniques, as they either defects within the LDH particles or separate phase(s) associated with LDHs. In contrast, samples prepared by coprecipitation especially synthesized under careful pH control and subsequently hydrothermal treated have high local order and good crystallinity (particle size). Our results show that both molecular level and macroscopic techniques are needed to assess the composition of LDHs, as the conventional PXRD and TEM analysis of LDHs failed to identify the many structural defects and/or amorphous phases.

  6. The Double Solid Reactant Method for modeling the release of trace elements from dissolving solid phases: I. Outline and limitations

    NASA Astrophysics Data System (ADS)

    Accornero, Marina; Marini, Luigi

    2008-10-01

    A Double Solid Reactant Method was elaborated from a suggestion of Marini (Geological sequestration of carbon dioxide: Thermodynamics, kinetics, and reaction path modeling. Developments in Geochemistry, Elsevier, Amsterdam, 2007) to simulate the release of trace elements during the progressive dissolution of solid phases. The method is based on the definition, for each dissolving solid, of both an entity whose thermodynamic and kinetic properties are known (either a pure mineral or a solid mixture) and a special reactant, that is, a material of known stoichiometry and unknown thermodynamic and kinetic properties. The special reactant is utilised to take into account the concentrations of trace elements in the dissolving solid phase. In this communication, the influence of several trace elements on the Δ G f o, Δ G r o and log K of the minerals considered by Lelli et al. (Environ Geol, 2007) and Accornero and Marini (Geobasi, 2007a; Proceedings of IMWA symposium, Cagliari, 27 31 May 2007b) was evaluated assuming ideal mixing in the solid state. These effects were found to be negligible for albite and the leucite latitic glass, limited for muscovites and chlorites, and slightly more important for apatites. These influences become progressively higher with increasing concentration of trace elements in these minerals. Based on these deviations in thermodynamic parameters, special reactants should not include oxide components with molar fractions higher than 0.003.

  7. Boronated monoclonal antibody conjugates for neutron capture therapy

    SciTech Connect

    Borg, D.C.; Elmore, J.J. Jr.; Ferrone, S.

    1986-01-01

    This paper describes the effectiveness of /sup 10/B-labeled monoclonal antibodies against Colo-38 human melanoma in vitro. The authors obtained high boron to antibody ratios while maintaining antibody activity by using dextran intermediate carriers to link /sup 10/B to the antibody. They developed a double cell quasi-competitive binding bioassay to minimize the effects of nonspecific binding of boronated complexes to cells. 1 fig., 2 tabs.

  8. Production of a broad specificity antibody for the development and validation of an optical SPR screening method for free and intracellular microcystins and nodularin in cyanobacteria cultures.

    PubMed

    Devlin, Shauna; Meneely, Julie P; Greer, Brett; Campbell, Katrina; Vasconcelos, Vitor; Elliott, Christopher T

    2014-05-01

    A highly sensitive broad specificity monoclonal antibody was produced and characterised for microcystin detection through the development of a rapid surface plasmon resonance (SPR) optical biosensor based immunoassay. The antibody displayed the following cross-reactivity: MC-LR 100%; MC-RR 108%; MC-YR 68%; MC-LA 69%; MC-LW 71%; MC-LF 68%; and Nodularin 94%. Microcystin-LR was covalently attached to a CM5 chip and with the monoclonal antibody was employed in a competitive 4 min injection assay to detect total microcystins in water samples below the WHO recommended limit (1 µg/L). A 'total microcystin' level was determined by measuring free and intracellular concentrations in cyanobacterial culture samples as this toxin is an endotoxin. Glass bead beating was used to lyse the cells as a rapid extraction procedure. This method was validated according to European Commission Decision 96/23/EC criteria. The method was proven to measure intracellular microcystin levels, the main source of the toxin, which often goes undetected by other analytical procedures and is advantageous in that it can be used for the monitoring of blooms to provide an early warning of toxicity. It was shown to be repeatable and reproducible, with recoveries from spiked samples ranging from 74 to 123%, and had % CVs below 10% for intra-assay analysis and 15% for inter-assay analysis. The detection capability of the assay was calculated as 0.5 ng/mL for extracellular toxins and 0.05 ng/mL for intracellular microcystins. A comparison of the SPR method with LC-MS/MS was achieved by testing six Microcystis aeruginosa cultures and this study yielded a correlation R(2) value of 0.9989. PMID:24720955

  9. Synthesis of [Zn-Al-CO 3] layered double hydroxides by a coprecipitation method under steady-state conditions

    NASA Astrophysics Data System (ADS)

    Chang, Z.; Evans, D. G.; Duan, X.; Vial, C.; Ghanbaja, J.; Prevot, V.; de Roy, M.; Forano, C.

    2005-09-01

    A continuous co-precipitation method under steady-state conditions has been investigated for the preparation of nanometer-size layered double hydroxide (LDH) particles using Zn 2Al(OH) 6(CO 3) 0.5·2H 2O as a prototype. The objective was to shorten the preparation time by working without an aging step, using a short and controlled residence time in order to maintain a constant supersaturation level in the reactor and constant particle properties in the exit stream over time. The effects of varying the operating conditions on the structural and textural properties of the LDHs have been studied, including total cation concentration, solvent, residence time, pH and intercalation anion. The products have been characterized using ICP, XRD, FTIR, BET, SEM and TEM. The LDHs prepared by the continuous coprecipitation method have a poorer crystallinity and lower crystallite sizes than those synthesized by the conventional batch method. The results have shown that increasing either cation concentration or the fraction of monoethylene glycol (MEG) in MEG/H 2O mixtures up to 80% (v/v) affect salt solubility and supersaturation, which gives rise to smaller crystallites, larger surface areas and more amorphous compounds. This increase is however limited by the precipitation of zinc and aluminum hydroxides occurring around a total cation concentration of 3.0×10 -1 M in pure water and 3.0×10 -2 M in H 2O/EtOH mixtures. Crystallite size increases with residence time, suggesting a precipitation process controlled by growth. Finally, the continuous coprecipitation method under steady-state conditions has been shown to be a promising alternative to the traditional coprecipitation technique in either pure water or mixed H 2O/MEG solvents.

  10. Antinuclear antibodies in domestic animals.

    PubMed

    Gershwin, Laurel J

    2005-06-01

    Antinuclear antibodies in domestic animal species have been commonly detected for many years, with the greatest frequency occurring in dogs as well as horses and cats. Most commonly, the assay used in diagnostic laboratories is indirect immunofluorescence on HEP-2 cells, similar to that used in human medicine, but with the exception that species-specific antiglobulin reagents are used instead of antihuman immunoglobulin. To a lesser extent, the Crithidia luciliae test for antibodies to double-stranded DNA has been used. Several research groups have used other assays. PMID:16014553

  11. [Targeted therapy by monoclonal antibodies].

    PubMed

    Ohnuma, Kei; Morimoto, Chikao

    2010-10-01

    Human monoclonal antibodies are virtually indispensable for immunotherapy of cancer, infectious diseases, autoimmune diseases, or organ transplantation. The hybridoma technique, developed by Georges Köhler and César Milstein in 1975, has been shown to be most and highly producible method for generating murine monoclonal antibodies. However, poor results were obtained when it was administered in human bodies. With development of biotechnology, human monoclonal antibodies have been manufactured with higher efficiency. A major hindrance of producing therapeutic human monoclonal antibodies is the lack of an appropriate strategy for determining and selecting the antibodies that would be effective in vivo. In this review, we give an overview of the present techniques on therapeutic monoclonal antibodies. PMID:20954327

  12. Antibodies as stratagems against cancer.

    PubMed

    Papageorgiou, Louis; Cuong, Nguyen Tien; Vlachakis, Dimitrios

    2016-06-21

    Antibodies have been in the frontline of anticancer research during the last few decades, since a number of different ways have been discovered to utilize them as parts or main components of anticancer drugs. Antibodies are used as the only component of some anticancer drugs, but they can also be conjugated with a variety of substances. Antibody engineering methods such as humanization, chimerization and Fc engineering are applied in order to modify their properties according to the requirements of anticancer drug application. Given the continuous advances in biology and informatics, the role of antibodies in anticancer treatment is expected to be prominent. PMID:26738941

  13. Precise characterization method of antibody-conjugated magnetic nanoparticles for pathogen detection using stuffer-free multiplex ligation-dependent probe amplification.

    PubMed

    Chung, Boram; Shin, Gi Won; Choi, Woong; Joo, Jinmyoung; Jeon, Sangmin; Jung, Gyoo Yeol

    2014-12-01

    Antibody-conjugated magnetic nanoparticles (Ab-MNPs) have potential in pathogen detection because they allow target cells to be easily separated from complex sample matrices. However, the sensitivity and specificity of pathogen capture by Ab-MNPs generally vary according to the types of MNPs, antibodies, and sample matrices, as well as preparation methods, including immobilization. Therefore, achieving a reproducible analysis utilizing Ab-MNPs as a pathogen detection method requires accurate characterization of Ab-MNP capture ability and standardization of all handling processes. In this study, we used high-resolution CE-single strand conformational polymorphism coupled with a stuffer-free multiplex ligation-dependent probe amplification system to characterize Ab-MNPs. The capture ability of Ab-MNPs targeting Salmonella enteritidis and nine pathogens, including S. enteritidis, was analyzed in phosphate buffer and milk. The effect of storage conditions on the stability of Ab-MNPs was also assessed. The results showed that the stuffer-free multiplex ligation-dependent probe amplification system has the potential to serve as a standard characterization method for Ab-MNPs. Moreover, the precise characterization of Ab-MNPs facilitated robust pathogen detection in various applications. PMID:25070923

  14. A sensitive multidimensional method for the detection, characterization, and quantification of trace free drug species in antibody-drug conjugate samples using mass spectral detection.

    PubMed

    Birdsall, Robert E; McCarthy, Sean M; Janin-Bussat, Marie Claire; Perez, Michel; Haeuw, Jean-François; Chen, Weibin; Beck, Alain

    2016-01-01

    Conjugation processes and stability studies associated with the production and shelf life of antibody-drug conjugates (ADCs) can result in free (non-conjugated) drug species. These free drug species can increase the risk to patients and reduce the efficacy of the ADC. Despite stringent purification steps, trace levels of free drug species may be present in formulated ADCs, reducing the therapeutic window. The reduction of sample preparation steps through the incorporation of multidimensional techniques has afforded analysts more efficient methods to assess trace drug species. Multidimensional methods coupling size-exclusion and reversed phase liquid chromatography with ultra-violet detection (SEC-RPLC/UV) have been reported, but offer limited sensitivity and can limit method optimization. The current study addresses these challenges with a multidimensional method that is specific, sensitive, and enables method control in both dimensions via coupling of an on-line solid phase extraction column to RPLC with mass spectral detection (SPE-RPLC/MS). The proposed method was evaluated using an antibody-fluorophore conjugate (AFC) as an ADC surrogate to brentuximab vedotin and its associated parent maleimide-val-cit-DSEA payload and the derived N-acetylcysteine adduct formed during the conjugation process. Assay sensitivity was found to be 2 orders more sensitive using MS detection in comparison to UV-based detection with a nominal limit of quantitation of 0.30 ng/mL (1.5 pg on-column). Free-drug species were present in an unadulterated ADC surrogate sample at concentrations below 7 ng/mL, levels not detectable by UV alone. The proposed SPE-RPLC/MS method provides a high degree of specificity and sensitivity in the assessment of trace free drug species and offers improved control over each dimension, enabling straightforward integration into existing or novel workflows. PMID:26651262

  15. A sensitive multidimensional method for the detection, characterization, and quantification of trace free drug species in antibody-drug conjugate samples using mass spectral detection

    PubMed Central

    Birdsall, Robert E.; McCarthy, Sean M.; Janin-Bussat, Marie Claire; Perez, Michel; Haeuw, Jean-François; Chen, Weibin; Beck, Alain

    2016-01-01

    abstract Conjugation processes and stability studies associated with the production and shelf life of antibody-drug conjugates (ADCs) can result in free (non-conjugated) drug species. These free drug species can increase the risk to patients and reduce the efficacy of the ADC. Despite stringent purification steps, trace levels of free drug species may be present in formulated ADCs, reducing the therapeutic window. The reduction of sample preparation steps through the incorporation of multidimensional techniques has afforded analysts more efficient methods to assess trace drug species. Multidimensional methods coupling size-exclusion and reversed phase liquid chromatography with ultra-violet detection (SEC-RPLC/UV) have been reported, but offer limited sensitivity and can limit method optimization. The current study addresses these challenges with a multidimensional method that is specific, sensitive, and enables method control in both dimensions via coupling of an on-line solid phase extraction column to RPLC with mass spectral detection (SPE-RPLC/MS). The proposed method was evaluated using an antibody-fluorophore conjugate (AFC) as an ADC surrogate to brentuximab vedotin and its associated parent maleimide-val-cit-DSEA payload and the derived N-acetylcysteine adduct formed during the conjugation process. Assay sensitivity was found to be 2 orders more sensitive using MS detection in comparison to UV-based detection with a nominal limit of quantitation of 0.30 ng/mL (1.5 pg on-column). Free-drug species were present in an unadulterated ADC surrogate sample at concentrations below 7 ng/mL, levels not detectable by UV alone. The proposed SPE-RPLC/MS method provides a high degree of specificity and sensitivity in the assessment of trace free drug species and offers improved control over each dimension, enabling straightforward integration into existing or novel workflows. PMID:26651262

  16. Homoclinic behaviors and chaotic motions of double layered viscoelastic nanoplates based on nonlocal theory and extended Melnikov method

    SciTech Connect

    Wang, Yu; Wang, Yi-Ze; Li, Feng-Ming

    2015-06-15

    The nonlinear dynamical equations are established for the double layered viscoelastic nanoplates (DLNP) subjected to in-plane excitation based on the nonlocal theory and von Kármán large deformation theory. The extended high dimensional homoclinic Melnikov method is employed to study the homoclinic phenomena and chaotic motions for the parametrically excited DLNP system. The criteria for the homoclinic transverse intersection for both the asynchronous and synchronous buckling cases are proposed. Lyapunov exponents and phase portraits are obtained to verify the Melnikov-type analysis. The influences of structural parameters on the transverse homoclinic orbits and homoclinic bifurcation sets are discussed for the two buckling cases. Some novel phenomena are observed in the investigation. It should be noticed that the nonlocal effect on the homoclinic behaviors and chaotic motions is quite remarkable. Hence, the small scale effect should be taken into account for homoclinic and chaotic analysis for nanostructures. It is significant that the nonlocal effect on the homoclinic phenomena for the asynchronous buckling case is quite different from that for the synchronous buckling case. Moreover, due to the van der Walls interaction between the layers, the nonlocal effect on the homoclinic behaviors and chaotic motions for high order mode is rather tiny under the asynchronous buckling condition.

  17. Analytic energy gradients for the coupled-cluster singles and doubles method with the density-fitting approximation.

    PubMed

    Bozkaya, Uğur; Sherrill, C David

    2016-05-01

    An efficient implementation is presented for analytic gradients of the coupled-cluster singles and doubles (CCSD) method with the density-fitting approximation, denoted DF-CCSD. Frozen core terms are also included. When applied to a set of alkanes, the DF-CCSD analytic gradients are significantly accelerated compared to conventional CCSD for larger molecules. The efficiency of our DF-CCSD algorithm arises from the acceleration of several different terms, which are designated as the "gradient terms": computation of particle density matrices (PDMs), generalized Fock-matrix (GFM), solution of the Z-vector equation, formation of the relaxed PDMs and GFM, back-transformation of PDMs and GFM to the atomic orbital (AO) basis, and evaluation of gradients in the AO basis. For the largest member of the alkane set (C10H22), the computational times for the gradient terms (with the cc-pVTZ basis set) are 2582.6 (CCSD) and 310.7 (DF-CCSD) min, respectively, a speed up of more than 8-folds. For gradient related terms, the DF approach avoids the usage of four-index electron repulsion integrals. Based on our previous study [U. Bozkaya, J. Chem. Phys. 141, 124108 (2014)], our formalism completely avoids construction or storage of the 4-index two-particle density matrix (TPDM), using instead 2- and 3-index TPDMs. The DF approach introduces negligible errors for equilibrium bond lengths and harmonic vibrational frequencies. PMID:27155621

  18. Analysis for chemical characterization of atmospheric aerosols application of X-ray microprobe system and double thin film method.

    PubMed

    Tohno, Susumu; Ma, Chang-Jin; Hayakawa, Shinjiro; Yamasaki, Satoshi; Kasahara, Mikio

    2006-09-01

    The X-ray microprobe system was applied to ultra trace characterization of single Kosa aerosols and non-Kosa aerosols simultaneously collected at Yasaka, Japan and TaeAnn, Korea. We demonstrated remarkable mass increase of heavy metals as well as soil components in individual particles during the Kosa event compared with the non-Kosa period at Yasaka. Backward trajectory analysis suggested that the Kosa samples were in the mixing state of mineral components and anthropogenic heavy elements. Double thin film method was applied to investigate the seasonal change of the mixing states of single sea-salt aerosols associated with chlorine loss due to the heterogeneous reactions between sea-salt particles and acidic gases. It was revealed that the percentages of both chloride-nitrate mixed particles and sulfate-nitrate ones were larger in wintertime than those in summertime and fraction of chloride-nitrate mixed particles increased with an increase of particle size. Comparison between the size-segregated bulk analysis and the single particle analysis demonstrated that remarkable chloride depletion occurred in coarse particles sampled on May in the former analysis, while chloride depletion in coarse particles was not marked in the latter analysis. The discrepancy attributes to the difference of the sampling time between both analyses because significant change of air mass route occurred during the bulk sampling after completion of the single particle sampling. PMID:16741794

  19. Homoclinic behaviors and chaotic motions of double layered viscoelastic nanoplates based on nonlocal theory and extended Melnikov method.

    PubMed

    Wang, Yu; Li, Feng-Ming; Wang, Yi-Ze

    2015-06-01

    The nonlinear dynamical equations are established for the double layered viscoelastic nanoplates (DLNP) subjected to in-plane excitation based on the nonlocal theory and von Kármán large deformation theory. The extended high dimensional homoclinic Melnikov method is employed to study the homoclinic phenomena and chaotic motions for the parametrically excited DLNP system. The criteria for the homoclinic transverse intersection for both the asynchronous and synchronous buckling cases are proposed. Lyapunov exponents and phase portraits are obtained to verify the Melnikov-type analysis. The influences of structural parameters on the transverse homoclinic orbits and homoclinic bifurcation sets are discussed for the two buckling cases. Some novel phenomena are observed in the investigation. It should be noticed that the nonlocal effect on the homoclinic behaviors and chaotic motions is quite remarkable. Hence, the small scale effect should be taken into account for homoclinic and chaotic analysis for nanostructures. It is significant that the nonlocal effect on the homoclinic phenomena for the asynchronous buckling case is quite different from that for the synchronous buckling case. Moreover, due to the van der Walls interaction between the layers, the nonlocal effect on the homoclinic behaviors and chaotic motions for high order mode is rather tiny under the asynchronous buckling condition. PMID:26117102

  20. Analytic energy gradients for the coupled-cluster singles and doubles method with the density-fitting approximation

    NASA Astrophysics Data System (ADS)

    Bozkaya, Uǧur; Sherrill, C. David

    2016-05-01

    An efficient implementation is presented for analytic gradients of the coupled-cluster singles and doubles (CCSD) method with the density-fitting approximation, denoted DF-CCSD. Frozen core terms are also included. When applied to a set of alkanes, the DF-CCSD analytic gradients are significantly accelerated compared to conventional CCSD for larger molecules. The efficiency of our DF-CCSD algorithm arises from the acceleration of several different terms, which are designated as the "gradient terms": computation of particle density matrices (PDMs), generalized Fock-matrix (GFM), solution of the Z-vector equation, formation of the relaxed PDMs and GFM, back-transformation of PDMs and GFM to the atomic orbital (AO) basis, and evaluation of gradients in the AO basis. For the largest member of the alkane set (C10H22), the computational times for the gradient terms (with the cc-pVTZ basis set) are 2582.6 (CCSD) and 310.7 (DF-CCSD) min, respectively, a speed up of more than 8-folds. For gradient related terms, the DF approach avoids the usage of four-index electron repulsion integrals. Based on our previous study [U. Bozkaya, J. Chem. Phys. 141, 124108 (2014)], our formalism completely avoids construction or storage of the 4-index two-particle density matrix (TPDM), using instead 2- and 3-index TPDMs. The DF approach introduces negligible errors for equilibrium bond lengths and harmonic vibrational frequencies.

  1. Antithyroid microsomal antibody

    MedlinePlus

    ... Thyroid antimicrosomal antibody; Antimicrosomal antibody; Microsomal antibody; Thyroid peroxidase antibody; TPOAb Images Blood test References Guber HA, Faraq AF. Evaluation of endocrine function. In: McPherson RA, Pincus MR, eds. Henry's Clinical ...

  2. Searching for Dual AGNs in Galaxy Mergers: Understanding Double-Peaked [O III] and Ultra Hard X-rays as Selection Method

    NASA Astrophysics Data System (ADS)

    McGurk, Rosalie C.; Max, Claire E.; Medling, Anne; Shields, Gregory A.

    2015-01-01

    When galaxies merge, gas accretes onto both central supermassive black holes. Thus, one expects to see close pairs of active galactic nuclei (AGNs), or dual AGNs, in a fraction of galaxy mergers. However, finding them remains a challenge. The presence of double-peaked [O III] or of ultra hard X-rays have been proposed as techniques to select dual AGNs efficiently. We studied a sample of double-peaked narrow [O III] emitting AGNs from SDSS DR7. By obtaining new and archival high spatial resolution images taken with the Keck 2 Laser Guide Star Adaptive Optics system and the near-infrared (IR) camera NIRC2, we showed that 30% of double-peaked [O III] emission line SDSS AGNs have two spatial components within a 3' radius. However, spatially resolved spectroscopy or X-ray observations are needed to confirm these galaxy pairs as systems containing two AGNs. We followed up these spatially-double candidate dual AGNs with integral field spectroscopy from Keck OSIRIS and Gemini GMOS and with long-slit spectroscopy from Keck NIRSPEC and Shane Kast Double Spectrograph. We find double-peaked emitters are caused sometimes by dual AGN and sometimes by outflows or narrow line kinematics. We also performed Chandra X-ray ACIS-S observations on 12 double-peaked candidate dual AGNs. Using our observations and 8 archival observations, we compare the distribution of X-ray photons to our spatially double near-IR images, measure X-ray luminosities and hardness ratios, and estimate column densities. By assessing what fraction of double-peaked emission line SDSS AGNs are true dual AGNs, we can better determine whether double-peaked [O III] is an efficient dual AGN indicator and constrain the statistics of dual AGNs. A second technique to find dual AGN is the detection of ultra hard X-rays by the Swift Burst Alert Telescope. We use CARMA observations to measure and map the CO(1-0) present in nearby ultra-hard X-ray Active Galactic Nuclei (AGNs) merging with either a quiescent companion

  3. Isolation and identification of Duck tembusu virus strain lH and development of latex-agglutination diagnostic method for rapid detection of antibodies.

    PubMed

    Wang, Quanxi; Wen, Yaping; Yifan Huang; Wu, Yijian; Cai, Yilong; Xu, Lihui; Wang, Changkang; Li, Ang; Wu, Baocheng; Chen, Jilong

    2014-12-01

    SUMMARY. An outbreak of egg-drop syndrome occurred on a Sheldrake duck farm in Longhai in Fujian Province, China, in 2012. The main clinical symptoms were sharply reduced egg production, crooked necks, and death. We isolated the virus from the sick ducks, identified it, and observed the histopathologic changes after viral infection. We detected viral RNA in the blood and feces of the infected ducks and developed a latex-agglutination diagnostic method to detect anti-Tembusu-virus antibodies. Our results show that the pathogenic virus is a Tembusu virus. The histopathologic changes included follicular cell degeneration and necrosis, follicular cavity filled with blood cells, massive necrosis in the brain, and degeneration and necrosis of the nerve and glial cells. When the transmission of the virus in the infected ducks was studied, the duck blood was positive for viral nucleic acid for up to 29 days, and the feces were positive for viral nucleic acid for up to 13 days. We successfully established a simple, rapid, and easy- to-use latex-agglutination diagnostic method for the detection of antibodies against duck Tembusu virus. PMID:25619007

  4. A Highly Selective and Sensitive Fluorescence Detection Method of Glyphosate Based on an Immune Reaction Strategy of Carbon Dot Labeled Antibody and Antigen Magnetic Beads.

    PubMed

    Wang, Duo; Lin, Bixia; Cao, Yujuan; Guo, Manli; Yu, Ying

    2016-08-01

    A sensitive fluorescence detection method for glyphosate (GLY) was established based on immune reaction. First, carbon dot labeled antibodies (lgG-CDs) which were able to specifically identify glyphosate were prepared with the environmentally friendly carbon dots (CDs) and glyphosate antibody (lgG). lgG-CDs could be used to in situ visualize the distribution of glyphosate in plant tissues. In order to eliminate the effects of excess lgG-CDs on the determination of GLY, antigen magnetic beads Fe3O4-GLY based on magnetic nanoparticles Fe3O4 and glyphosate were constructed and utilized to couple with the excess lgG-CDs. After magnetic separation to remove antigen magnetic beads, there was a linear relationship between the fluorescence intensity of lgG-CDs and the logarithmic concentration of glyphosate in the range of 0.01-80 μg/mL with a detection limit of 8 ng/mL. The method was used for the detection of glyphosate in Pearl River water, tea, and soil samples with satisfactory recovery ratio between 87.4% and 103.7%. PMID:27403652

  5. A facile method for the construction of covalently cross-linked layered double hydroxides layer-by-layer films: Enhanced stability and delayed release of guests

    NASA Astrophysics Data System (ADS)

    Li, Yulong; An, Qi; Hu, Yingmo; Luan, Xinglong; Zhang, Qian; Zhang, Tianhang; Zhang, Yihe

    2015-07-01

    Stable composite films that contain layered double hydroxide (LDH) are appealing materials but are also difficult to prepare. We report here a facile strategy for the fabrication of covalently cross-linked layer-by-layer multilayers that incorporate LDH. The films were first prepared using the traditional LbL method based on non-covalent interactions, followed by infiltration of a photoactive small molecule DAS. UV light was then used to cross-link the multilayers. The stability of the cross-linked film was remarkably enhanced. Furthermore, the release profile of incorporated molecules from layered double hydroxide was significantly delayed.

  6. Deposition of nanometric double layers Ru/Au, Ru/Pd, and Pd/Au onto CdZnTe by the electroless method

    NASA Astrophysics Data System (ADS)

    Zheng, Q.; Dierre, F.; Corregidor, V.; Fernández-Ruiz, R.; Crocco, J.; Bensalah, H.; Alves, E.; Diéguez, E.

    2012-11-01

    Nanometric double layer properties of different metal-semiconductor-metal (MSM) depositions onto CdZnTe produced by electroless method are investigated. The mechanisms of the deposition are discussed and the theoretical chemical equations implied in the process are presented. The solutions for different time of deposition and the deposited layers are analysed by TXRF to test the proposed reactions. RBS was used to determine the thickness, the depth profiles and the composition of the layers deposited at the surface. This work showed the feasibility of depositing nanometric double layers using electroless technique.

  7. Fast and simple epidemiological typing of Pseudomonas aeruginosa using the double-locus sequence typing (DLST) method.

    PubMed

    Basset, P; Blanc, D S

    2014-06-01

    Although the molecular typing of Pseudomonas aeruginosa is important to understand the local epidemiology of this opportunistic pathogen, it remains challenging. Our aim was to develop a simple typing method based on the sequencing of two highly variable loci. Single-strand sequencing of three highly variable loci (ms172, ms217, and oprD) was performed on a collection of 282 isolates recovered between 1994 and 2007 (from patients and the environment). As expected, the resolution of each locus alone [number of types (NT) = 35-64; index of discrimination (ID) = 0.816-0.964] was lower than the combination of two loci (NT = 78-97; ID = 0.966-0.971). As each pairwise combination of loci gave similar results, we selected the most robust combination with ms172 [reverse; R] and ms217 [R] to constitute the double-locus sequence typing (DLST) scheme for P. aeruginosa. This combination gave: (i) a complete genotype for 276/282 isolates (typability of 98%), (ii) 86 different types, and (iii) an ID of 0.968. Analysis of multiple isolates from the same patients or taps showed that DLST genotypes are generally stable over a period of several months. The high typability, discriminatory power, and ease of use of the proposed DLST scheme makes it a method of choice for local epidemiological analyses of P. aeruginosa. Moreover, the possibility to give unambiguous definition of types allowed to develop an Internet database ( http://www.dlst.org ) accessible by all. PMID:24326699

  8. Elevated antimeasles antibody titre: An association in autoimmune encephalitis

    PubMed Central

    Chandra, S. R.; Issac, Thomas Gregor; Philip, Mariamma; Krishnan, A.

    2015-01-01

    Introduction: Autoimmune encephalitis is a group of treatable noninfective encephalitic disorders with great clinical implications. They have a close resemblance to prion disease and some slow virus infections. We report the presence of significant titers of antimeasles antibody in some of our patients with autoimmune encephalitis resulting in diagnostic and therapeutic problems. Patients and Methods: Patients seen by us in the last 4 years with high titers (1:625 dilution) cerebrospinal fiuid (CSF) antimeasles antibody positivity were reviewed retrospectively. The data collected were assessed using SPSS- Statistical Package for Social Sciences Version 15.0 (IBM corporation) software. The groups which showed elevated antimeasles antibody titers but did not have other parameters suggestive of subacute sclerosing panencephalitis (Group 2) were segregated and compared with those who had the typical features (Group 1) using Fisher's Exact Test. Results: There were 33 patients with antimeasles antibody in CSF. Group 1 had 27 and Group 2 had 6 patients. Group 1 had lower age, cognitive dysfunction, slow myoclonus, less generalized tonic-clonic seizures, and focal seizures. Group 2 patients belonged to the higher age, had significant psychosis (P = 0.02), incontinence of bowel and bladder (P = 0.0001). Slow myoclonus was significant in the first group (P = 0.028), and weakness was significant in the second group (P = 0.028) and double incontinence in the second group (P = 0.0001). Magnetic resonance imaging showed significant gray matter and cerebellar involvement in Group 2 P = 0.005 and P = 0.028, respectively. Conclusions: Patients who show significant titers of antimeasles antibodies in the CSF but belonging to older age group with psychosis, generalized tonic-clonic seizures, double incontinence, focal myoclonus, and electroencephalographic and imaging noncorroborative need to be investigated for autoimmune encephalitis in view of the great prognostic and therapeutic

  9. Efficacy of interference screw and double-docking methods using palmaris longus and GraftJacket for medial collateral ligament reconstruction of the elbow.

    PubMed

    Furukawa, Keizo; Pichora, Jamie; Steinmann, Scott; Faber, Kenneth J; Johnson, James A; King, Graham J W

    2007-01-01

    Single-strand elbow medial collateral ligament reconstruction strength was evaluated by use of double-docking and interference screw methods with either a palmaris longus tendon or GraftJacket as the reconstruction material. Thirteen upper extremities were mounted in 90 degrees of valgus orientation and subjected to cyclic valgus loading that increased progressively until failure occurred. The double-docking reconstructions outperformed the interference screw reconstructions (P < .05), whereas the palmaris longus and GraftJacket performed comparably (P > .05). The favorable initial strength of the GraftJacket make it a potentially viable alternative to the use of autogenous palmaris longus tendons; however, further studies are required to evaluate graft strength during healing. The clinical use of the double-docking technique of single-strand medial collateral ligament reconstruction should be considered because of its simplicity and initial strength. PMID:17368922

  10. A Double-Coil TMS Method to Assess Corticospinal Excitability Changes at a Near-Simultaneous Time in the Two Hands during Movement Preparation

    PubMed Central

    Wilhelm, Emmanuelle; Quoilin, Caroline; Petitjean, Charlotte; Duque, Julie

    2016-01-01

    Background: Many previous transcranial magnetic stimulation (TMS) studies have investigated corticospinal excitability changes occurring when choosing which hand to use for an action, one of the most frequent decision people make in daily life. So far, these studies have applied single-pulse TMS eliciting motor-evoked potential (MEP) in one hand when this hand is either selected or non-selected. Using such method, hand choices were shown to entail the operation of two inhibitory mechanisms, suppressing MEPs in the targeted hand either when it is non-selected (competition resolution, CR) or selected (impulse control, IC). However, an important limitation of this “Single-Coil” method is that MEPs are elicited in selected and non-selected conditions during separate trials and thus those two settings may not be completely comparable. Moreover, a more important problem is that MEPs are computed in relation to the movement of different hands. The goal of the present study was to test a “Double-Coil” method to evaluate IC and CR preceding the same hand responses by applying Double-Coil TMS over the two primary motor cortices (M1) at a near-simultaneous time (1 ms inter-pulse interval). Methods: MEPs were obtained in the left (MEPLEFT) and right (MEPRIGHT) hands while subjects chose between left and right hand key-presses in blocks using a Single-Coil or a Double-Coil method; in the latter blocks, TMS was either applied over left M1 first (TMSLRM1 group, n = 12) or right M1 first (TMSRLM1 group, n = 12). Results: MEPLEFT were suppressed preceding both left (IC) and right (CR) hand responses whereas MEPRIGHT were only suppressed preceding left (CR) but not right (IC) hand responses. This result was observed regardless of whether Single-Coil or Double-Coil TMS was applied in the two subject groups. However, in the TMSLRM1 group, the MEP suppression was attenuated in Double-Coil compared to Single-Coil blocks for both IC and CR, when probed with MEPLEFT (elicited by

  11. Molecular-specific urokinase antibodies

    NASA Technical Reports Server (NTRS)

    Atassi, M. Zouhair (Inventor); Morrison, Dennis R. (Inventor)

    2009-01-01

    Antibodies have been developed against the different molecular forms of urokinase using synthetic peptides as immunogens. The peptides were synthesized specifically to represent those regions of the urokinase molecules which are exposed in the three-dimensional configuration of the molecule and are uniquely homologous to urokinase. Antibodies are directed against the lysine 158-isoleucine 159 peptide bond which is cleaved during activation from the single-chain (ScuPA) form to the bioactive double chain (54 KDa and 33 KDa) forms of urokinase and against the lysine 135 lysine 136 bond that is cleaved in the process of removing the alpha-chain from the 54 KDa form to produce the 33 KDa form of urokinase. These antibodies enable the direct measurement of the different molecular forms of urokinase from small samples of conditioned medium harvested from cell cultures.

  12. Combined DNA extraction and antibody elution from filter papers for the assessment of malaria transmission intensity in epidemiological studies

    PubMed Central

    2013-01-01

    Background Informing and evaluating malaria control efforts relies on knowledge of local transmission dynamics. Serological and molecular tools have demonstrated great sensitivity to quantify transmission intensity in low endemic settings where the sensitivity of traditional methods is limited. Filter paper blood spots are commonly used a source of both DNA and antibodies. To enhance the operational practicability of malaria surveys, a method is presented for combined DNA extraction and antibody elution. Methods Filter paper blood spots were collected as part of a large cross-sectional survey in the Kenyan highlands. DNA was extracted using a saponin/chelex method. The eluate of the first wash during the DNA extraction process was used for antibody detection and compared with previously validated antibody elution procedures. Antibody elution efficiency was assessed by total IgG ELISA for malaria antigens apical membrane antigen-1 (AMA-1) and merozoite-surface protein-1 (MSP-142). The sensitivity of nested 18S rRNA and cytochrome b PCR assays and the impact of doubling filter paper material for PCR sensitivity were determined. The distribution of cell material and antibodies throughout filter paper blood spots were examined using luminescent and fluorescent reporter assays. Results Antibody levels measured after the combined antibody/DNA extraction technique were strongly correlated to those measured after standard antibody elution (p < 0.0001). Antibody levels for both AMA-1 and MSP-142 were generally slightly lower (11.3-21.4%) but age-seroprevalence patterns were indistinguishable. The proportion of parasite positive samples ranged from 12.9% to 19.2% in the different PCR assays. Despite strong agreement between outcomes of different PCR assays, none of the assays detected all parasite-positive individuals. For all assays doubling filter paper material for DNA extraction increased sensitivity. The concentration of cell and antibody material was not

  13. Universal method for synthesis of artificial gel antibodies by the imprinting approach combined with a unique electrophoresis technique for detection of minute structural differences of proteins, viruses and cells (bacteria). Ib. Gel antibodies against proteins (hemoglobins).

    PubMed

    Takátsy, Anikó; Végvári, Akos; Hjertén, Stellan; Kilár, Ferenc

    2007-07-01

    Using the molecular imprinting approach, we have shown that polyacrylamide-based artificial antibodies against human and bovine hemoglobin have a very high selectivity, as revealed by the free-zone electrophoresis in a revolving capillary. By the same technique we have previously synthesized gel antibodies not only against proteins but also against viruses and bacteria. The synthesis is thus universal, i.e., it has the great advantage of not requiring a modification - or only a slight one - for each particular antigen. The combination synthesis of artificial gel antibodies and electrophoretic analysis reveals small discrepancies in shape and chemical composition not only of proteins, as shown here and in paper Ia, but also of viruses and bacteria, to be illustrated in papers II and III in this series. Upon rehydration, the freeze-dried gel antibodies, selective for human hemoglobin, regain their selectivity. The gel antibodies can repeatedly be used following the removal of the antigen (protein in this study) from the complex gel antibody/antigen by an SDS washing or an enzymatic degradation. PMID:17476715

  14. The double solid reactant method: II. An application to the shallow groundwaters of the Porto Plain, Vulcano Island (Italy)

    NASA Astrophysics Data System (ADS)

    Lelli, Matteo; Cioni, Roberto; Marini, Luigi

    2008-11-01

    This paper presents an example of application of the double solid reactant method (DSRM) of Accornero and Marini (Environmental Geology, 2007a), an effective way for modeling the fate of several dissolved trace elements during water-rock interaction. The EQ3/6 software package was used for simulating the irreversible water-rock mass transfer accompanying the generation of the groundwaters of the Porto Plain shallow aquifer, starting from a degassed diluted crateric steam condensate. Reaction path modeling was performed in reaction progress mode and under closed-system conditions. The simulations assumed: (1) bulk dissolution (i.e., without any constraint on the kinetics of dissolution/precipitation reactions) of a single solid phase, a leucite-latitic glass, and (2) precipitation of amorphous silica, barite, alunite, jarosite, anhydrite, kaolinite, a solid mixture of smectites, fluorite, a solid mixture of hydroxides, illite-K, a solid mixture of saponites, a solid mixture of trigonal carbonates and a solid mixture of orthorhombic carbonates. Analytical concentrations of major chemical elements and several trace elements (Cr, Mn, Fe, Ni, Cu, Zn, As, Sr and Ba) in groundwaters were satisfactorily reproduced. In addition to these simulations, similar runs for a rhyolite, a latite and a trachyte permitted to calculate major oxide contents for the authigenic paragenesis which are comparable, to a first approximation, with the corresponding data measured for local altered rocks belonging to the silicic, advanced argillic and intermediate argillic alteration facies. The important role played by both the solid mixture of trigonal carbonates as sequestrator of Mn, Zn, Cu and Ni and the solid mixture of orthorhombic carbonates as scavenger of Sr and Ba is emphasized.

  15. Application of a risk analysis method to different technologies for producing a monoclonal antibody employed in hepatitis B vaccine manufacturing.

    PubMed

    Milá, Lorely; Valdés, Rodolfo; Tamayo, Andrés; Padilla, Sigifredo; Ferro, Williams

    2012-03-01

    CB.Hep-1 monoclonal antibody (mAb) is used for a recombinant Hepatitis B vaccine manufacturing, which is included in a worldwide vaccination program against Hepatitis B disease. The use of this mAb as immunoligand has been addressed into one of the most efficient steps of active pharmaceutical ingredient purification process. Regarding this, Quality Risk Management (QRM) provides an excellent framework for the risk management use in pharmaceutical manufacturing and quality decision-making applications. Consequently, this study sought applying a prospective risk analysis methodology Failure Mode Effects Analysis (FMEA) as QRM tool for analyzing different CB.Hep-1 mAb manufacturing technologies. As main conclusions FMEA was successfully used to assess risks associated with potential problems in CB.Hep-1 mAb manufacturing processes. The severity and occurrence of risks analysis evidenced that the percentage of very high severe risks ranged 31.0-38.7% of all risks and the huge majority of risks have a very low occurrence level (61.9-83.3%) in all assessed technologies. Finally, additive Risk Priority Number, was descending ordered as follow: transgenic plants (2636), ascites (2577), transgenic animals (2046) and hollow fiber bioreactors (1654), which also corroborated that in vitro technology, should be the technology of choice for CB.Hep-1 mAb manufacturing in terms of risks and mAb molecule quality. PMID:22285820

  16. Double peak-induced distance error in short-time-Fourier-transform-Brillouin optical time domain reflectometers event detection and the recovery method.

    PubMed

    Yu, Yifei; Luo, Linqing; Li, Bo; Guo, Linfeng; Yan, Jize; Soga, Kenichi

    2015-10-01

    The measured distance error caused by double peaks in the BOTDRs (Brillouin optical time domain reflectometers) system is a kind of Brillouin scattering spectrum (BSS) deformation, discussed and simulated for the first time in the paper, to the best of the authors' knowledge. Double peak, as a kind of Brillouin spectrum deformation, is important in the enhancement of spatial resolution, measurement accuracy, and crack detection. Due to the variances of the peak powers of the BSS along the fiber, the measured starting point of a step-shape frequency transition region is shifted and results in distance errors. Zero-padded short-time-Fourier-transform (STFT) can restore the transition-induced double peaks in the asymmetric and deformed BSS, thus offering more accurate and quicker measurements than the conventional Lorentz-fitting method. The recovering method based on the double-peak detection and corresponding BSS deformation can be applied to calculate the real starting point, which can improve the distance accuracy of the STFT-based BOTDR system. PMID:26479653

  17. [Optimization of ELISA and immunoblot methods for the detection of IgG antibodies against old world hantaviruses in wild rodents].

    PubMed

    Polat, Ceylan; Karataş, Ahmet; Sözen, Mustafa; Matur, Ferhat; Abacıoğlu, Hakan; Öktem, Mehmet Ali

    2016-04-01

    Hantaviruses infect humans via inhalation of viral particles in infected rodents' secretions such as saliva, urine and faeces or via direct contact with infected rodents. The rodent species that are known as the carriers of Dobrava (DOBV), Puumala (PUUV), Saaremaa (SAAV), Tula (TULV) and Seoul (SEOV) viruses are found in our country. The presence of specific antibodies against hantaviruses have been demonstrated in rodents collected from Black Sea and Aegean Regions of Turkey in 2004 for the first time. The first hantavirus-related hemorrhagic fever with renal syndrome (HFRS) cases were reported in Black Sea region in 2009. The determination of the hantavirus prevalence in wild life and rodent populations in the field is crucial for the information about hantavirus-related cases and to clarify the state of risk. There is no commercial product optimized for the screening of rodent serum samples in terms of HFRS agents like DOBV and PUUV that are widely seen in Eurasia as well as Turkey. In this study, the antigens belonging to the commercial enzyme-linked immunoassay (ELISA) and immunoblot tests that are produced for the screening of human sera were used for the development of antibody screening tests against hantavirus in rodent sera and were optimized. The most appropriate serum and conjugate dilutions were determined for the optimization of ELISA (Anti-Hantavirus Pool ELISA; Euroimmun, Germany) and immunoblot (Euroline Anti-Hanta Profile 1 strips; Euroimmun, Germany) methods. Optimized ELISA method was used for the screening and optimized immunoblot method was used for the confirmation. A total of 84 wild rodent sera that belonged to Apodemus and Microtus species were evaluated with this procedure and the cut-off value, sensitivity and specificity of optimized ELISA method were determined. For the optimization of ELISA 1/50, 1/100 and 1/200 serum dilutions and 1/10.000, 1/20.000 and 1/40.000 conjugate dilutions were tested. For the optimization of immunoblot, 1

  18. Antibody Engineering and Therapeutics

    PubMed Central

    Almagro, Juan Carlos; Gilliland, Gary L; Breden, Felix; Scott, Jamie K; Sok, Devin; Pauthner, Matthias; Reichert, Janice M; Helguera, Gustavo; Andrabi, Raiees; Mabry, Robert; Bléry, Mathieu; Voss, James E; Laurén, Juha; Abuqayyas, Lubna; Barghorn, Stefan; Ben-Jacob, Eshel; Crowe, James E; Huston, James S; Johnston, Stephen Albert; Krauland, Eric; Lund-Johansen, Fridtjof; Marasco, Wayne A; Parren, Paul WHI; Xu, Kai Y

    2014-01-01

    The 24th Antibody Engineering & Therapeutics meeting brought together a broad range of participants who were updated on the latest advances in antibody research and development. Organized by IBC Life Sciences, the gathering is the annual meeting of The Antibody Society, which serves as the scientific sponsor. Preconference workshops on 3D modeling and delineation of clonal lineages were featured, and the conference included sessions on a wide variety of topics relevant to researchers, including systems biology; antibody deep sequencing and repertoires; the effects of antibody gene variation and usage on antibody response; directed evolution; knowledge-based design; antibodies in a complex environment; polyreactive antibodies and polyspecificity; the interface between antibody therapy and cellular immunity in cancer; antibodies in cardiometabolic medicine; antibody pharmacokinetics, distribution and off-target toxicity; optimizing antibody formats for immunotherapy; polyclonals, oligoclonals and bispecifics; antibody discovery platforms; and antibody-drug conjugates. PMID:24589717

  19. [The significance of antiphospholipid antibodies].

    PubMed

    Fojtík, Z

    2004-04-01

    Antiphospholipid antibodies (APLA) present very heterogeneous groups of antibodies which can significantly influence processes on different levels of coagulation cascade depending on effects of phospholipid surfaces on blood coagulation. This usually leads to a particular level of thrombophylia. Clinical syndrome accompanying positive APLA, such as antiphospholipid syndrome, was defined by clinical and laboratory symptoms. This clinical syndrome can be a primary syndrome, if other disorders with ability to induce generation of antibodies can be excluded, or a secondary syndrome. The most often in cases of systemic tissue disease. APLA can be divided according to the presence of lupus anticoagulant and anticardiolipin antibodies. According to a definition lupus anticoagulants are antibodies able to inhibit and prolong in vitro one or more blood clotting processes dependent on phospholipid surfaces. Anticardiolipin antibodies are antibodies measured by ELISA method with cardiolipin used as an antibody. Findings show that some APLA are directed against proteins bound to phospholipid surfaces. Main cofactor proteins include beta 2-GPI and prothrombin. Because of their heterogeneous specificity, APLA are directed against negative phospholipids or proteins bound to phospholipid surfaces and have important pathophysiology role in development of antiphospholipid syndrome. PMID:15214303

  20. Combustion method for assay of biological materials labeled with carbon-14 or tritium, or double-labeled

    NASA Technical Reports Server (NTRS)

    Huebner, L. G.; Kisieleski, W. E.

    1969-01-01

    Dry catalytic combustion at high temperatures is used for assaying biological materials labeled carbon-14 and tritium, or double-labeled. A modified oxygen-flask technique is combined with standard vacuum-line techniques and includes convenience of direct in-vial collection of final combustion products, giving quantitative recovery of tritium and carbon-14.

  1. Development and evaluation of an antibody ELISA for sarcoptic mange in sheep and a comparison with the skin-scraping method.

    PubMed

    Rodríguez-Cadenas, F; Carbajal-González, M T; Fregeneda-Grandes, J M; Aller-Gancedo, J M; Huntley, J F; Rojo-Vázquez, F A

    2010-08-01

    In this work an indirect ELISA for detecting serum-specific IgG antibodies in sheep was developed using a crude saline extract from Sarcoptes scabiei var. ovis mites and then the repeatability of the ELISA outcomes was estimated. Subsequently, its diagnostic accuracy was evaluated by Receiver Operating Characteristics (ROC) analysis using a sample collected from the entire sheep population of western Castile and Leon region in Spain, and then compared with that of the skin-scraping method. The reference method used was a combination of clinical examination, skin-scraping analysis and epidemiological surveys, but it introduced selection and probably information biases. Furthermore, we attempted to identify biological factors useful to predict the sensitivity or specificity of the ELISA as determined by comparison with the reference method. Additionally, conventional latent-class analysis [Hui, S.L., Walter, S.D., 1980. Estimating the error rates of diagnostic tests. Biometrics 36, 167-171] was also used to estimate accuracy parameters. The between-run coefficient of variation (CV) for a standard serum was 8.8% and the within-run CV 4.3%. No significant deviation between the OD% means and strength positive correlation between the OD% values (r=0.98) were found for the results from two different batches of antigen. When compared to the reference method, the Area Under the ROC curve (AUC) for the reference population was 0.967 (95% CI: 0.949-0.985) for the ELISA and 0.915 (95% CI: 0.863-0.968) for the skin-scraping method. By logistic regression analysis, one explanatory biological factor-result to the skin-scraping method-and four explanatory biological factors-Tyroglyphidae individual status, Trichophyton verrucosum individual status, Oestrus ovis status of the flock and presence of adjacent animals with a clinical disease neighbour to S. scabiei infection-were found for diagnostic sensitivity and specificity of the ELISA, respectively, although this depended on the

  2. A rapid on-line method for mass spectrometric confirmation of a cysteine-conjugated antibody-drug-conjugate structure using multidimensional chromatography

    PubMed Central

    Birdsall, Robert E; Shion, Henry; Kotch, Frank W; Xu, April; Porter, Thomas J; Chen, Weibin

    2015-01-01

    Cysteine-conjugated antibody-drug conjugates (ADCs) are manufactured using controlled partial reduction and conjugation chemistry with drug payloads that typically occur in intervals of 0, 2, 4, 6, and 8. Control of heterogeneity is of particular importance to the quality of ADC product because drug loading and distribution can affect the safety and efficacy of the ADC. Liquid chromatography ultra-violet (LC-UV)-based methods can be used to acquire the drug distribution profiles of cysteine-conjugated ADCs when analyzed using hydrophobic interaction chromatography (HIC). However, alternative analysis techniques are often required for structural identification when conjugated drugs do not possess discrete ultra-violet absorbance properties for precise assessment of the drug-to-antibody ratio (DAR). In this study, multidimensional chromatography was used as an efficient method for combining non-compatible techniques, such as HIC, with analysis by mass spectrometry (LC/LC/QTOF-MS) for rapid on-line structural elucidation of species observed in HIC distribution profiles of cysteine-conjugated ADCs. The methodology was tested using an IgG1 mAb modified by cysteine conjugation with a non-toxic drug mimic. Structural elucidation of peaks observed in the HIC analysis (1st dimension) were successfully identified based on their unique sub-unit masses via mass spectrometry techniques once dissociation occurred under denaturing reversed phase conditions (2nd dimension). Upon identification, the DAR values were determined to be 2.83, 4.44, and 5.97 for 3 drug load levels (low-, medium-, and high-loaded ADC batches), respectively, based on relative abundance from the LC-UV data. This work demonstrates that multidimensional chromatography coupled with MS, provides an efficient approach for on-line biotherapeutic characterization to ensure ADC product quality. PMID:26305867

  3. A rapid on-line method for mass spectrometric confirmation of a cysteine-conjugated antibody-drug-conjugate structure using multidimensional chromatography.

    PubMed

    Birdsall, Robert E; Shion, Henry; Kotch, Frank W; Xu, April; Porter, Thomas J; Chen, Weibin

    2015-01-01

    Cysteine-conjugated antibody-drug conjugates (ADCs) are manufactured using controlled partial reduction and conjugation chemistry with drug payloads that typically occur in intervals of 0, 2, 4, 6, and 8. Control of heterogeneity is of particular importance to the quality of ADC product because drug loading and distribution can affect the safety and efficacy of the ADC. Liquid chromatography ultra-violet (LC-UV)-based methods can be used to acquire the drug distribution profiles of cysteine-conjugated ADCs when analyzed using hydrophobic interaction chromatography (HIC). However, alternative analysis techniques are often required for structural identification when conjugated drugs do not possess discrete ultra-violet absorbance properties for precise assessment of the drug-to-antibody ratio (DAR). In this study, multidimensional chromatography was used as an efficient method for combining non-compatible techniques, such as HIC, with analysis by mass spectrometry (LC/LC/QTOF-MS) for rapid on-line structural elucidation of species observed in HIC distribution profiles of cysteine-conjugated ADCs. The methodology was tested using an IgG1 mAb modified by cysteine conjugation with a non-toxic drug mimic. Structural elucidation of peaks observed in the HIC analysis (1(st) dimension) were successfully identified based on their unique sub-unit masses via mass spectrometry techniques once dissociation occurred under denaturing reversed phase conditions (2(nd) dimension). Upon identification, the DAR values were determined to be 2.83, 4.44, and 5.97 for 3 drug load levels (low-, medium-, and high-loaded ADC batches), respectively, based on relative abundance from the LC-UV data. This work demonstrates that multidimensional chromatography coupled with MS, provides an efficient approach for on-line biotherapeutic characterization to ensure ADC product quality. PMID:26305867

  4. Investigating the Quantitative Structure-Activity Relationships for Antibody Recognition of Two Immunoassays for Polycyclic Aromatic Hydrocarbons by Multiple Regression Methods

    PubMed Central

    Zhang, Yan-Feng; Zhang, Li; Gao, Zhi-Xian; Dai, Shu-Gui

    2012-01-01

    Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous contaminants found in the environment. Immunoassays represent useful analytical methods to complement traditional analytical procedures for PAHs. Cross-reactivity (CR) is a very useful character to evaluate the extent of cross-reaction of a cross-reactant in immunoreactions and immunoassays. The quantitative relationships between the molecular properties and the CR of PAHs were established by stepwise multiple linear regression, principal component regression and partial least square regression, using the data of two commercial enzyme-linked immunosorbent assay (ELISA) kits. The objective is to find the most important molecular properties that affect the CR, and predict the CR by multiple regression methods. The results show that the physicochemical, electronic and topological properties of the PAH molecules have an integrated effect on the CR properties for the two ELISAs, among which molar solubility (Sm) and valence molecular connectivity index (3χv) are the most important factors. The obtained regression equations for RisC kit are all statistically significant (p < 0.005) and show satisfactory ability for predicting CR values, while equations for RaPID kit are all not significant (p > 0.05) and not suitable for predicting. It is probably because that the RisC immunoassay employs a monoclonal antibody, while the RaPID kit is based on polyclonal antibody. Considering the important effect of solubility on the CR values, cross-reaction potential (CRP) is calculated and used as a complement of CR for evaluation of cross-reactions in immunoassays. Only the compounds with both high CR and high CRP can cause intense cross-reactions in immunoassays. PMID:23012547

  5. Standardization of a method to detect bovine sperm-bound anti-sperm antibodies by flow cytometry

    PubMed Central

    Sardoy, MC; Anderson, DE; George, A; Wilkerson, ME; Skinner, S; Ferrer, MS

    2012-01-01

    The objectives were to standardize some methodological and analytical aspects of a direct technique to detect sperm-bound anti-sperm antibodies (ASAs) in bovine semen using flow cytometry, including the effects of pre-fixation of sperm membranes with formalin buffer solution and inclusion of dead cells in the analysis. Fourteen Angus bulls, including ASA-positive (experimentally induced ASAs) and 10 reproductively normal ASA-negative bulls, were used. Fixation of sperm membranes had no significant effect on the percentage of IgG- or IgA-bound spermatozoa detected by flow cytometry. However, including dead cells in the analysis increased the percentage of IgG-bound spermatozoa in fixed (live and dead 18.6 ± 9.7% and live 1.3 ± 0.5%; median ± SEM) and non-fixed samples (live and dead 18.8 ± 9.2%, live 1.5 ± 0.6%; P = 0.0029), as well as IgA-bound spermatozoa in fixed (live and dead 16.3 ± 6.4%, live 0.3 ± 0.5%) and non-fixed samples (live and dead 21.4 ± 4.6%, live 1.0 ± 0.5%; P = 0.0041) in semen from ASA-negative bulls. Intra-sample, intra-assay and inter-assay coefficients of variation (CV) were 0.8, 4.6 and 5.3%, respectively, for determination of sperm-bound IgG, and were 2.8, 8.4 and 40.3% for determination of sperm-bound IgA. Despite the high inter-assay CV for IgA determination, all ASA-positive bulls consistently had high percentages of IgA-bound spermatozoa. Flow cytometry correctly identified ASA-positive bulls. Confocal laser microscopy confirmed binding of ASAs to sperm heads and cytoplasmic droplets, and less frequently to midpieces and principal piece. In conclusion, although fixation was not necessary, dead cells should be excluded from the analysis, since ejaculates with a large proportion of dead cells can yield false-positive results. Flow cytometry was accurate and reliable for detection of sperm-bound IgG and IgA and discrimination between ASA-positive and ASA-negative bulls. PMID:22925638

  6. Complete-mouth rehabilitation using a 3D printing technique and the CAD/CAM double scanning method: A clinical report.

    PubMed

    Joo, Han-Sung; Park, Sang-Won; Yun, Kwi-Dug; Lim, Hyun-Pil

    2016-07-01

    According to evolving computer-aided design/computer-aided manufacturing (CAD/CAM) technology, ceramic materials such as zirconia can be used to create fixed dental prostheses for partial removable dental prostheses. Since 3D printing technology was introduced a few years ago, dental applications of this technique have gradually increased. This clinical report presents a complete-mouth rehabilitation using 3D printing and the CAD/CAM double-scanning method. PMID:26946918

  7. ELECTROMAGNETISM, OPTICS, ACOUSTICS, HEAT TRANSFER, CLASSICAL MECHANICS, AND FLUID DYNAMICS: Double Symplectic Eigenfunction Expansion Method of Free Vibration of Rectangular Thin Plates

    NASA Astrophysics Data System (ADS)

    Wang, Hua; Alatancang; Huang, Jun-Jie

    2009-12-01

    The free vibration problem of rectangular thin plates is rewritten as a new upper triangular matrix differential system. For the associated operator matrix, we find that the two diagonal block operators are Hamiltonian. Moreover, the existence and completeness of normed symplectic orthogonal eigenfunction systems of these two block operators are demonstrated. Based on the completeness, the general solution of the free vibration of rectangular thin plates is given by double symplectic eigenfunction expansion method.

  8. Efficiency of sex pre-selection of spermatozoa by albumin separation method evaluated by double-labelled fluorescence in-situ hybridization.

    PubMed

    Chen, M J; Guu, H F; Ho, E S

    1997-09-01

    To evaluate the separation efficiency of Ericsson's two- and three-layer albumin separation methods, semen samples from 21 healthy males were studied. Seven patients already had two or more sons, another seven had two or more daughters and the other seven had primary infertility due to female factors. The semen samples were divided into three aliquots: one remained unprocessed initially, the other two aliquots went through two- and three-layer albumin separation methods respectively. All samples were then stained with X-Y double staining probes. In each group, four or five samples were processed at room temperature, and two or three at body temperature (37 degrees C). The labelling efficiency of X-Y double staining probe was over 99%. The X:Y sperm ratios were even in the original samples. The ratios of the X and Y spermatozoa were altered slightly but significantly after the two-layer (P < or = 0.05) or the three-layer (P < or = 0.005) separation. The alterations occurred only at room temperature. The X spermatozoa increased and the Y spermatozoa decreased, both to a small degree of difference (1.4-3.5%). Double fluorescence in-situ hybridization analysis therefore showed that albumin separation methods do not enrich Y spermatozoa. PMID:9363707

  9. Prediction of Antibody Epitopes.

    PubMed

    Nielsen, Morten; Marcatili, Paolo

    2015-01-01

    Antibodies recognize their cognate antigens in a precise and effective way. In order to do so, they target regions of the antigenic molecules that have specific features such as large exposed areas, presence of charged or polar atoms, specific secondary structure elements, and lack of similarity to self-proteins. Given the sequence or the structure of a protein of interest, several methods exploit such features to predict the residues that are more likely to be recognized by an immunoglobulin. Here, we present two methods (BepiPred and DiscoTope) to predict linear and discontinuous antibody epitopes from the sequence and/or the three-dimensional structure of a target protein. PMID:26424260

  10. Setting of Methods for Analysis of Mucosal Antibodies in Seminal and Vaginal Fluids of HIV Seropositive Subjects from Cambodian and Italian Cohorts

    PubMed Central

    Scotti, Lorenza; Diomede, Lorenzo; Ngyen, Marie; Nouhin, Janin; DeSantis, Lucia; Zambon, Antonella; Ferrari, Davide; Gallotta, Giulia; Corrao, Giovanni; Pancino, Gianfranco; Lopalco, Lucia

    2010-01-01

    Background Genital mucosae play a key role in protection from STD and HIV infection, due to their involvement in both horizontal and vertical disease transmission. High variability of published observations concerning IgA isolation and quantification underlies the strong requirement of specific methods able to maximize investigation on HIV-specific IgA. Methodology Genital fluids from 109 subjects, including male and female cohorts from Italy and Cambodia, were collected, aliquoted and processed with different techniques, to assess optimal conditions maximizing mucosal antibody recovery. Three sampling techniques, up to sixteen preservation conditions, six ELISA methods and four purifications protocols were compared. Principal Findings The optimal method here described took advantage of Weck-Cel sampling of female mucosal fluids. Immediate processing of genital fluids, with the addition of antibiotics and EDTA, improved recovery of vaginal IgA, while the triple addition of EDTA, antibiotics and protease inhibitors provided the highest amount of seminal IgA. Due to low amount of IgA in mucosal fluids, a high sensitive sandwich ELISA assay was set; sensitivity was enhanced by milk-based overcoating buffer and by a two-step biotin-streptavidin signal amplification. Indeed, commercial antisera to detect human immunoglobulins showed weak cross-reactivity to different antibody types. Three-step affinity purification provided reproducible immunoglobulin recovery from genital specimens, while conventional immuno-affinity IgA purification was found poorly manageable. Affinity columns were suitable to isolate mucosal IgA, which are ten-fold less concentrated than IgG in genital specimens, and provided effective separation of IgA monomers, dimers, and J-chains. Jacalin-bound resin successfully separated IgA1 from IgA2 subfraction. Conclusions/Significance Specific, effective and reliable methods to study local immunity are key items in understanding host mucosal response. The

  11. Technology Review of Nondestructive Methods for Examination of Water Intrusion Areas on Hanford’s Double-Shell Waste Tanks

    SciTech Connect

    Watkins, Michael L.; Pardini, Allan F.

    2008-05-09

    Under a contract with CH2M Hill Hanford Group, Inc., PNNL has performed a review of the NDE technology and methods for examination of the concrete dome structure of Hanford’s double-shell tanks. The objective was to provide a matrix of methodologies that could be evaluated based on applicability, ease of deployment, and results that could provide information that could be used in the ongoing structural analysis of the tank dome. PNNL performed a technology evaluation with the objective of providing a critical literature review for all applicable technologies based on constraints provided by CH2M HILL. These constraints were not mandatory, but were desired. These constraints included performing the evaluation without removing any soil from the top of the tank, or if necessary, requesting that the hole diameter needed to gain access to evaluate the top of the tank structure to be no greater than approximately 12-in. in diameter. PNNL did not address the details of statistical sampling requirements as they depend on an unspecified risk tolerance. PNNL considered these during the technology evaluation and have reported the results in the remainder of this document. Many of the basic approaches to concrete inspection that were reviewed in previous efforts are still in use. These include electromagnetic, acoustic, radiographic, etc. The primary improvements in these tools have focused on providing quantitative image reconstruction, thus providing inspectors and analysts with three-dimensional data sets that allow for operator visualization of relevant abnormalities and analytical integration into structural performance models. Available instruments, such as radar used for bridge deck inspections, rely on post-processing algorithms and do not provide real-time visualization. Commercially available equipment only provides qualitative indications of relative concrete damage. It cannot be used as direct input for structural analysis to assess fitness for use and if

  12. Finite Element Method Simulation of Double-T Type Vibrators for Flatly Supported Piezoelectric Vibratory Gyro-Sensor

    NASA Astrophysics Data System (ADS)

    Kikuchi, Takayuki; Tomikawa, Yoshiro; Soma, Takao

    2001-05-01

    In this study we deal with a flexural vibrating resonator for flatly supported vibratory gyro-sensors in parallel with the rotating plane. This type of gyro-sensor uses two vibration modes in only one plane of single crystal quartz, and therefore can be constructed to be very stable under operating conditions. The double-T type vibrator that has two T-type resonating arms and two resonant arms is designed for improving the quality factor Q of both driving and detecting vibration modes for this vibratory gyro-sensor. The result of finite element analysis of the double-T type vibrators shows the small influence of its vibration mode with the support at the center of gravity of the vibrator.

  13. Trifunctional antibody ertumaxomab

    PubMed Central

    Diermeier-Daucher, Simone; Ortmann, Olaf; Buchholz, Stefan; Brockhoff, Gero

    2012-01-01

    Background: The trifunctional antibody ertumaxomab bivalently targets the human epidermal growth factor receptor 2 (Her2) on epithelial (tumor) cells and the T cell specific CD3 antigen, and its Fc region is selectively recognized by Fcγ type I/III receptor-positive immune cells. As a trifunctional immunoglobulin, ertumaxomab therefore not only targets Her2 on cancer cells, but also triggers immunological effector mechanisms mediated by T and accessory cells (e.g., macrophages, dendritic cells, natural killer cells). Whether molecular effects, however, might contribute to the cellular antitumor efficiency of ertumaxomab are largely unknown. Methods: Potential molecular effects of ertumaxomab on Her2-overexpressing BT474 and SK-BR-3 breast cancer cells were evaluated. The dissociation constant Kd of ertumaxomab was calculated from titration curves that were recorded by flow cytometry. Treatment-induced changes in Her2 homodimerization were determined by flow cytometric fluorescence resonance energy transfer measurements on a cell-by-cell basis. Potential activation / deactivation of Her2, ERK1/2, AKT and STAT3 were analyzed by western blotting, Immunochemistry and immunofluorescent cell staining. Results: The Kd of ertumaxomab for Her2-binding was determined at 265 nM and the ertumaxomab binding epitope was found to not overlap with that of the therapeutic anti-Her2 monoclonal antibodies trastuzumab and pertuzumab. Ertumaxomab caused an increase in Her2 phosphorylation at higher antibody concentrations, but changed neither the rate of Her2-homodimerization /-phosphorylation nor the activation state of key downstream signaling proteins analyzed. Conclusions: The unique mode of action of ertumaxomab, which relies more on activation of immune-mediated mechanisms against tumor cells compared with currently available therapeutic antibodies for breast cancer treatment, suggests that modular or sequential treatment with the trifunctional bivalent antibody might complement

  14. Serum herpes simplex antibodies

    MedlinePlus

    ... gov/ency/article/003352.htm Serum herpes simplex antibodies To use the sharing features on this page, please enable JavaScript. Serum herpes simplex antibodies is a blood test that looks for antibodies ...

  15. Antibodies against mumps virus component proteins.

    PubMed

    Matsubara, Keita; Iwata, Satoshi; Nakayama, Tetsuo

    2012-08-01

    The neutralization (NT) test is regarded as the most reliable method for detection of protective antibodies, but is labor-intensive and time consuming. Enzyme-linked immunosorbent assay (EIA) is frequently used in sero-epidemiological studies because of its simplicity and ease of use. In this study, immunofluorescent (IF) antibodies against nucleocapsid (N), fusion (F), and hemagglutinin-neuraminidase (HN) proteins were investigated in comparison with NT and EIA antibodies. The antibody against N protein was dominant in serum samples obtained from patients with a previous history of mumps infection. Titers of antibodies against F and HN proteins were very low. Many serum samples were positive for EIA but negative for NT, and no significant correlation was noted between NT and EIA antibodies. Among the three component proteins, correlation of EIA and IF antibodies with N protein was relatively good. After vaccination with mumps vaccine, EIA positivity was closely related to the IF antibodies against N protein, and after vaccination NT-positive sera became positive for IF antibodies against F and HN proteins. IF antibodies against F and HN proteins were considered to have a strong association with NT antibodies, and those against N protein were considered to have a strong association with EIA antibodies. PMID:22215227

  16. A Facile Method to Prepare Double-Layer Isoporous Hollow Fiber Membrane by In Situ Hydrogen Bond Formation in the Spinning Line.

    PubMed

    Noor, Nazia; Koll, Joachim; Radjabian, Maryam; Abetz, Clarissa; Abetz, Volker

    2016-03-01

    A double-layer hollow fiber is fabricated where an isoporous surface of polystyrene-block-poly(4-vinylpyridine) is fixed on a support layer by co-extrusion. Due to the sulfonation of the support layer material, delamination of the two layers is suppressed without increasing the number of subsequent processing steps for isoporous composite membrane formation. Electron microscope-energy-dispersive X-ray spectroscopy images unveil the existence of a high sulfur concentration in the interfacial region by which in-process H-bond formation between the layers is evidenced. For the very first time, our study reports a facile method to fabricate a sturdy isoporous double-layer hollow fiber. PMID:26685710

  17. The detection and discovery of glycan motifs in biological samples using lectins and antibodies: new methods and opportunities

    PubMed Central

    Tang, Huiyuan; Hsueh, Peter; Kletter, Doron; Bern, Marshall; Haab, Brian

    2015-01-01

    Recent research is uncovering unexpected ways that glycans contribute to biology, as well as new strategies for combatting disease using approaches involving glycans. To make full use of glycans for clinical applications, we need more detailed information on the location, nature, and dynamics of glycan expression in vivo. Such studies require the use of specimens acquired directly from patients. Effective studies of clinical specimens require low-volume assays, high precision measurements, and the ability to process many samples. Assays using affinity reagents—lectins and glycan-binding antibodies—can meet these requirements, but further developments are needed to make the methods routine and effective. Recent advances in the use of glycan-binding proteins could meet that need. The advances involve improved determination of specificity using glycan arrays; the availability of databases for mining and analyzing glycan array data; lectin engineering methods; and the ability to quantitatively interpret lectin measurements. Here we describe many of the challenges and opportunities involved in the application of these new approaches to the study of biological samples. The new tools hold promise for developing methods to improve the outcomes of patients afflicted with diseases characterized by aberrant glycan expression. PMID:25727148

  18. Cell force mapping using a double-sided micropillar array based on the moiré fringe method

    NASA Astrophysics Data System (ADS)

    Zhang, F.; Anderson, S.; Zheng, X.; Roberts, E.; Qiu, Y.; Liao, R.; Zhang, X.

    2014-07-01

    The mapping of traction forces is crucial to understanding the means by which cells regulate their behavior and physiological function to adapt to and communicate with their local microenvironment. To this end, polymeric micropillar arrays have been used for measuring cell traction force. However, the small scale of the micropillar deflections induced by cell traction forces results in highly inefficient force analyses using conventional optical approaches; in many cases, cell forces may be below the limits of detection achieved using conventional microscopy. To address these limitations, the moiré phenomenon has been leveraged as a visualization tool for cell force mapping due to its inherent magnification effect and capacity for whole-field force measurements. This Letter reports an optomechanical cell force sensor, namely, a double-sided micropillar array (DMPA) made of poly(dimethylsiloxane), on which one side is employed to support cultured living cells while the opposing side serves as a reference pattern for generating moiré patterns. The distance between the two sides, which is a crucial parameter influencing moiré pattern contrast, is predetermined during fabrication using theoretical calculations based on the Talbot effect that aim to optimize contrast. Herein, double-sided micropillar arrays were validated by mapping mouse embryo fibroblast contraction forces and the resulting force maps compared to conventional microscopy image analyses as the reference standard. The DMPA-based approach precludes the requirement for aligning two independent periodic substrates, improves moiré contrast, and enables efficient moiré pattern generation. Furthermore, the double-sided structure readily allows for the integration of moiré-based cell force mapping into microfabricated cell culture environments or lab-on-a-chip devices.

  19. Study of Method for Designing the Power and the Capacitance of Fuel Cells and Electric Double-Layer Capacitors of Hybrid Railway Vehicle

    NASA Astrophysics Data System (ADS)

    Takizawa, Kenji; Kondo, Keiichiro

    A hybrid railway traction system with fuel cells (FCs) and electric double layer-capacitors (EDLCs) is discussed in this paper. This system can save FC costs and absorb the regenerative energy. A method for designing FCs and EDLCs on the basis of the output power and capacitance, respectively, has not been reported, even though their design is one of the most important technical issues encountered in the design of hybrid railway vehicles. Such design method is presented along with a train load profile and an energy management strategy. The design results obtained using the proposed method are verified by performing numerical simulations of a running train. These results reveal that the proposed method for designing the EDLCs and FCs on the basis of the capacitance and power, respectively, and by using a method for controlling the EDLC voltage is sufficiently effective in designing efficient EDLCs and FCs of hybrid railway traction systems.

  20. Multiplexed evaluation of serum and CSF pharmacokinetics of brain-targeting single-domain antibodies using a NanoLC-SRM-ILIS method.

    PubMed

    Haqqani, Arsalan S; Caram-Salas, Nadia; Ding, Wen; Brunette, Eric; Delaney, Christie E; Baumann, Ewa; Boileau, Eve; Stanimirovic, Danica

    2013-05-01

    FC5 and FC44 are single-domain antibodies (VHHs), selected by functional panning of phage-display llama VHH library for their ability to internalize human brain endothelial cells (BEC) and to transmigrate the in vitro BBB model. Quantification of brain delivery of FC5 and FC44 in vivo was challenging using classical methods because of their short plasma half-life and their loss of functionality with radioactive labeling. A highly sensitive (detection limit <2 ng/mL) and specific SRM-ILIS method to detect and quantify unlabeled VHHs in multiplexed assays was developed and applied to comparatively evaluate brain delivery of FC5 and FC44, and two control VHHs, EG2 and A20.1. FC5 and FC44 compared to control VHHs demonstrated significantly (p < 0.01) enhanced transport (50-100-fold) across rat in vitro BBB model as well as in vivo brain targeting assessed by optical imaging. The multiplexed SRM-ILIS analyses of plasma and CSF levels of codosed VHHs demonstrated that while all 4 VHHs have similar blood pharmacokinetics, only FC5 and FC44 show elevated CSF levels, suggesting that they are potential novel carriers for delivery of drugs and macromolecules across the BBB. PMID:23150993

  1. Evaluation of a commercial ELISA for H5 low pathogenic avian influenza virus antibody detection in duck sera using Bayesian methods.

    PubMed

    Schmitz, Audrey; Le Bras, Marie-Odile; Guillemoto, Carole; Pierre, Isabelle; Rose, Nicolas; Bougeard, Stéphanie; Jestin, Véronique

    2013-10-01

    Following the emergence of highly pathogenic avian influenza (AI), active surveillance of infections due to the H5 and H7 subtypes in poultry has increased and been made compulsory in Europe since 2002, by means of annual serological surveys using the haemagglutination inhibition (HI) test. Domestic anseriforms, particularly ducks and geese, are more frequently infected by H5 low pathogenic AI virus, often subclinically, and represent a threat for other terrestrial poultry. 1783 sera, mainly from ducks, have been used to evaluate and compare a commercial ELISA kit detecting H5 antibodies with the currently recommended HI test. Different approaches to calculating specificity and sensitivity have been used, including the original Bayesian method. Results were similar when data were analyzed at the individual and batch levels, and when using different methods of calculation. However, results showed that H5 ELISA had both a higher sensitivity and a lower specificity than the HI test. Given that sensitivity is the most important factor for a screening test, H5 ELISA could therefore be recommended for AI surveillance, followed in cases of positivity by molecular tests aimed at detecting the virus gene. PMID:23727545

  2. Transfer of proteins from gels to diazobenzyloxymethyl-paper and detection with antisera: a method for studying antibody specificity and antigen structure.

    PubMed Central

    Renart, J; Reiser, J; Stark, G R

    1979-01-01

    We describe a rapid and very sensitive method for detecting proteins as antigens after their separation in polyacrylamide/agarose composite gels, with or without sodium dodecyl sulfate. The polyacrylamide matrix is crosslinked with a reagent that can be cleaved with periodate or alkali to facilitate transfer of the protein bands to diazobenzyloxymethyl-paper, where they are coupled covalently. Specific proteins are detected by autoradiography after sequential incubation with unfractionated, unlabeled specific antiserum and 125I-labeled protein A from Staphylococcus aureus. Antibody and protein A can be removed with urea and 2-mercaptoethanol, and the same paper can be probed again with a different antiserum. An antiserum specific for the simian virus 40 virion proteins VP3 and VP2 has been prepared; it does not crossreact with VP1, as demonstrated by this method. An antiserum raised in rabbits against simian virus 40-transformed rabbit kidney cells is shown to be directed primarily against a periodate-sensitive moiety present in tumor (T) antigen from infected or transformed cells, whereas an antiserum raised in rabbits against large T antigen purified from lytically infected monkey kidney cells by electrophoresis in the presence of sodium dodecyl sulfate [Lane, D.P. & Robbins, A.K. (1978) Virology 87, 182-193] is directed primarily against determinants that are not sensitive to periodate. Images PMID:91164

  3. Selection of antibodies from synthetic antibody libraries.

    PubMed

    Harel Inbar, Noa; Benhar, Itai

    2012-10-15

    More than 2 dozen years had passed since the field of antibody engineering was established, with the first reports of bacterial [1-3] and mammalian cells [4] expression of recombinant antibody fragments, and in that time a lot of effort was dedicated to the development of efficient technological means, intended to assist in the creation of therapeutic monoclonal antibodies (mAbs). Research focus was given to two intertwined technological aspects: the selection platform and the recombinant antibody repertoires. In accordance with these areas of interest, it is the goal of this chapter to describe the various selection tools and antibody libraries existing, with emphasis on the later, and their applications. This chapter gives a far from exhaustive, subjective "historic account" of the field, describing the selection platforms, the different formats of antibody repertoires and the applications of both for selecting recombinant antibodies. Several excellent books provide detailed protocols for constructing antibody libraries and selecting antibodies from those libraries [5-13]. Such books may guide a newcomer to the field in the fine details of antibody engineering. We would like to offer advice to the novice: although seemingly simple, effective library construction and antibody isolation provide best benefits in the hands of professionals. It is an art as much as it is science. PMID:22244834

  4. Comparative Study of Two Methods of Orthogonal Double-Pulse Laser-Induced Breakdown Spectroscopy of Aluminum

    NASA Astrophysics Data System (ADS)

    Safi, A.; Bahreini, M.; Tavassoli, S. H.

    2016-03-01

    Double-pulse laser induced breakdown spectroscopy (DP-LIBS) of aluminum sample is studied experimentally in orthogonal configuration in air. In this configuration, two schemes of reheating and pre-ablation are examined and the results are compared with single pulse one. The effect of delay time between two laser pulses on emission line intensities of plasma is investigated. Some of the parameters that have been involved in different mechanism of signal enhancement such as plasma temperature, sample heating effects, atmospheric effects, and modification of the ablation dynamics are more discussed. Investigation of the effect of laser pulse energy on emission line intensities in single pulse LIBS experiment demonstrate that because of saturation effects the intensities will not increase necessarily by increasing the laser pulse energy. Moreover, the results show that the electron temperature and rate of mass removal in orthogonal configuration of DP-LIBS is higher than that of single pulse with the same total energy. It is suggested that for correct comparison between single and double pulse results, the optimum pulse energy in single pulse should be considered. Overall, our results demonstrate that under optimized conditions the signal enhancement is much more in pre-ablation configuration than re-heating configuration.

  5. Drug delivery system for an anticancer agent, chlorogenate-Zn/Al-layered double hydroxide nanohybrid synthesised using direct co-precipitation and ion exchange methods

    SciTech Connect

    Barahuie, Farahnaz; Hussein, Mohd Zobir; Arulselvan, Palanisamy; Fakurazi, Sharida; Zainal, Zulkarnain

    2014-09-15

    A nano-structured drug-inorganic clay hybrid involving an active anticancer compound, which is chlorogenic acid (CA) intercalated into Zn/Al-layered double hydroxide, has been assembled via ion-exchange and co-precipitation methods to form a nanohybrid CZAE (a chlorogenic acid-Zn/Al nanohybrid synthesised using an ion-exchange method) and CZAC (a chlorogenic acid-Zn/Al nanohybrid synthesised using a direct method), respectively. The X-ray diffraction (XRD) results confirmed that the CA-LDH had a hybrid structure in which the anionic chlorogenate is arranged between the interlayers as a horizontal monolayer at 90 and 20° angles from the x axis for CZAE and CZAC, respectively. Both nanohybrids have the properties of mesoporous materials. The high loading percentage of chlorogenic acid (approximately 43.2% for CZAE and 45.3% for CZAC) with basal spacings of 11.7 and 12.6 Å for CZAE and CZAC, respectively, corroborates the successful intercalation of chlorogenic acid into the interlayer gallery of layered double hydroxides. Free chlorogenic acid and the synthesised nanocomposites (CZAE, CZAC) were assessed for their cytotoxicity against various cancer cells. The Fourier transform infrared data supported the formation of both nanohybrids, and a thermal analysis showed that the nanohybrids are more thermally stable than their counterparts. The chlorogenate shows a sustained release, and the release rate of chlorogenate from CZAE and CZAC nanohybrids at pH 7.4 is remarkably lower than that at pH 4.8 due to their different release mechanisms. The release rate of chlorogenate from both nanohybrids can be described as pseudo-second order. The present investigation revealed the potential of the nanohybrids to enhance the in vitro anti-tumour effect of chlorogenic acid in liver and lung cancer cells in vitro. - Highlights: • We intercalated chlorogenic into Zn/Al-layered double hydroxide by ion-exchange and coprecipitation methods. • The two methods gave nanocomposites

  6. Quantification of Five Clinically Important Amino Acids by HPLC-Triple TOF™ 5600 Based on Pre-column Double Derivatization Method.

    PubMed

    Deng, Shuang; Scott, David; Garg, Uttam

    2016-01-01

    Phenylalanine, tyrosine, glycine, cystine, and phosphoethanolamine are commonly measured amino acids in various physiological fluids to diagnose or follow-up various inborn errors of metabolism. The gold standard method for the amino acids quantitation has been ion exchange chromatography with ninhydrin post-column derivatization. However, this method is very laborious and time consuming. In recent years, liquid-chromatography mass spectrometry is being increasingly used for the assay of amino acids. Pre-column butyl derivatization with reverse phase chromatography has been widely used for mass spectrometry analysis of amino acids. Phosphoethanolamine is not butylated and cannot be measured by this method. Nevertheless, phosphoethanolamine can be dansyl-derivatized using dansyl chloride. We developed a double derivatization method by using butanol and dansyl chloride to derivatize carboxylic and amino groups separately, and then combining the derivatives to simultaneously measure these five amino acids using TOF-MS scan. Stable isotope-labeled internal standards were used. PMID:26602116

  7. Interaction analysis of HIV-1 antibody 2G12 and Man9GlcNAc2 ligand: Theoretical calculations by fragment molecular orbital and MD methods

    NASA Astrophysics Data System (ADS)

    Koyama, Yuka; Ueno-Noto, Kaori; Takano, Keiko

    2013-07-01

    In HIV-1 infection, human antibody 2G12 is capable of recognizing the high-mannose glycans on the HIV-1 surface glycoprotein, gp120. To investigate the ligand binding mechanisms of antibody 2G12 with glycans aiming for the contribution to the medications, we carried out classical molecular dynamics (MD) simulations and ab initio fragment molecular orbital (FMO) calculations on the antibody 2G12 complex with its high-mannose ligand. We found that Mannose D1 of the ligand had the largest binding affinity with the antibody, which was well consistent with experimental reports. Furthermore, significant roles of Mannose 4 and 4‧ in the ligand binding were theoretically indicated.

  8. Subzero temperature chromatography and top-down mass spectrometry for protein higher-order structure characterization: method validation and application to therapeutic antibodies.

    PubMed

    Pan, Jingxi; Zhang, Suping; Parker, Carol E; Borchers, Christoph H

    2014-09-17

    Characterization of the higher-order structure and structural dynamics of proteins is crucial for in-depth understanding of their functions. Amide hydrogen/deuterium exchange (HDX), monitored by mass spectrometry (MS), is now a popular technique for measuring protein higher-order structural changes. Although the proteolysis-based HDX-MS approach is most commonly used, the "top-down" approach, which fragments intact proteins directly using electron-based dissociation, is becoming an important alternative and has several advantages. However, the commonly used top-down strategies are direct-infusion based and thus can only be used with volatile buffers. This has meant that the "top-down" approach could not be used for studying proteins under physiological conditions-the very conditions which are often very important for preserving a protein's native structure and function. More complex proteins such as those with disulfide bonds present another challenge. Therefore, there is significant interest in developing novel top-down HDX methods that are applicable to all types of protein samples. In this paper, we show how top-down electron capture dissociation and subzero temperature HPLC can be combined and used for this purpose. This method keeps the back-exchange level as low as 2% and has no limitations in terms of protein type and sample solution conditions. Close to single-residue level protein structural information can be generated. The new method is validated through comparison with NMR data using calmodulin as a model protein. Its capability of determining structural changes in therapeutic antibodies (Herceptin) is also demonstrated. PMID:25152011

  9. Detection of Legionella species in sewage and ocean water by polymerase chain reaction, direct fluorescent-antibody, and plate culture methods.

    PubMed

    Palmer, C J; Tsai, Y L; Paszko-Kolva, C; Mayer, C; Sangermano, L R

    1993-11-01

    Legionella spp. are ubiquitous in most environmental water sources; however, sewage treatment plants have not been examined as potential environmental reservoirs for these bacteria. This study used polymerase chain reaction, direct fluorescent-antibody staining, and culture methods to examine raw and treated sewage, ocean-receiving waters, and nearshore coastal environments for the presence of Legionella pneumophila and other Legionella spp. The study concluded that Legionella spp. are present in all phases of sewage treatment and that population numbers do not significantly decline through the treatment process. Ocean-receiving waters located 5 miles offshore, where the treated sewage is discharged, were found to contain Legionella spp., but ocean water between the discharge site and coastal bathing beaches was negative. This suggests that the Legionella spp. from the ocean discharge site were not reaching the nearshore beach waters. A flood control channel and river that entered the ocean were found to contain Legionella spp., and a nearby beach swimming area was also found to be positive, suggesting that land runoff from the flood control channel and river were the source of the Legionella spp. in the beach water samples that tested positive. PMID:8285669

  10. Detection of Legionella species in sewage and ocean water by polymerase chain reaction, direct fluorescent-antibody, and plate culture methods.

    PubMed Central

    Palmer, C J; Tsai, Y L; Paszko-Kolva, C; Mayer, C; Sangermano, L R

    1993-01-01

    Legionella spp. are ubiquitous in most environmental water sources; however, sewage treatment plants have not been examined as potential environmental reservoirs for these bacteria. This study used polymerase chain reaction, direct fluorescent-antibody staining, and culture methods to examine raw and treated sewage, ocean-receiving waters, and nearshore coastal environments for the presence of Legionella pneumophila and other Legionella spp. The study concluded that Legionella spp. are present in all phases of sewage treatment and that population numbers do not significantly decline through the treatment process. Ocean-receiving waters located 5 miles offshore, where the treated sewage is discharged, were found to contain Legionella spp., but ocean water between the discharge site and coastal bathing beaches was negative. This suggests that the Legionella spp. from the ocean discharge site were not reaching the nearshore beach waters. A flood control channel and river that entered the ocean were found to contain Legionella spp., and a nearby beach swimming area was also found to be positive, suggesting that land runoff from the flood control channel and river were the source of the Legionella spp. in the beach water samples that tested positive. Images PMID:8285669

  11. Speeding up the solution of the Bethe-Salpeter equation by a double-grid method and Wannier interpolation

    NASA Astrophysics Data System (ADS)

    Kammerlander, David; Botti, Silvana; Marques, Miguel A. L.; Marini, Andrea; Attaccalite, Claudio

    2012-09-01

    The Bethe-Salpeter equation is a widely used approach to describe optical excitations in bulk semiconductors. It leads to spectra that are in very good agreement with experiment, but the price to pay for such accuracy is a very high computational burden. One of the main bottlenecks is the large number of k points required to obtain converged spectra. In order to circumvent this problem we propose a strategy to solve the Bethe-Salpeter equation based on a double-grid technique coupled to a Wannier interpolation of the Kohn-Sham band structure. This strategy is then benchmarked for a particularly difficult case, the calculation of the absorption spectrum of GaAs, and for the well-studied case of Si. The considerable gains observed in these cases fully validate our approach, and open the way for the application of the Bethe-Salpeter equation to large and complex systems.

  12. HANFORD DOUBLE SHELL TANK THERMAL AND SEISMIC PROJECT BUCKLING EVALUATION METHODS AND RESULTS FOR THE PRIMARY TANKS

    SciTech Connect

    MACKEY TC; JOHNSON KI; DEIBLER JE; PILLI SP; RINKER MW; KARRI NK

    2009-01-14

    This report documents a detailed buckling evaluation of the primary tanks in the Hanford double-shell waste tanks (DSTs), which is part of a comprehensive structural review for the Double-Shell Tank Integrity Project. This work also provides information on tank integrity that specifically responds to concerns raised by the Office of Environment, Safety, and Health (ES&H) Oversight (EH-22) during a review of work performed on the double-shell tank farms and the operation of the aging waste facility (AWF) primary tank ventilation system. The current buckling review focuses on the following tasks: (1) Evaluate the potential for progressive anchor bolt failure and the appropriateness of the safety factors that were used for evaluating local and global buckling. The analysis will specifically answer the following questions: (a) Can the EH-22 scenario develop if the vacuum is limited to -6.6-inch water gage (w.g.) by a relief valve? (b) What is the appropriate factor of safety required to protect against buckling if the EH-22 scenario can develop? (c) What is the appropriate factor of safety required to protect against buckling if the EH-22 scenario cannot develop? (2) Develop influence functions to estimate the axial stresses in the primary tanks for all reasonable combinations of tank loads based on detailed finite element analysis. The analysis must account for the variation in design details and operating conditions between the different DSTs. The analysis must also address the imperfection sensitivity of the primary tank to buckling. (3) Perform a detailed buckling analysis to determine the maximum allowable differential pressure for each of the DST primary tanks at the current specified limits on waste temperature, height, and specific gravity. Based on the concrete anchor bolt loads analysis and the small deformations that are predicted at the unfactored limits on vacuum and axial loads, it is very unlikely that the EH-22 scenario (i.e., progressive anchor bolt

  13. Reactions of organoaluminum compounds with acetylene as a method for the synthesis of aliphatic derivatives with a z-disubstituted double bond

    SciTech Connect

    Andreeva, N.I.; Kuchin, A.V.; Tolstikov, G.A.

    1985-11-01

    This paper develops a method for the synthesis of aliphatic compounds with a Z-disubstituted double bond, which are important synthons for the preparation of such natural products as insect pheromones, aromatic principles, etc. In the carbalumination reaction of acetylene Z-alkenyldialkylaluminums are formed selectively. A-Alkenyldialkylaluminums are highly reactive and can readily be converted into Z-allyl alcohols and their ethers, and into Z-iodovinyl derivatives. By the reactions of vinyl organoaluminum compounds with the complex CH/sub 3/COClhaAlCl/sub 3/ E-conjugated ketones were obtained.

  14. Emerging double helical nanostructures

    NASA Astrophysics Data System (ADS)

    Zhao, Meng-Qiang; Zhang, Qiang; Tian, Gui-Li; Wei, Fei

    2014-07-01

    As one of the most important and land-mark structures found in nature, a double helix consists of two congruent single helices with the same axis or a translation along the axis. This double helical structure renders the deoxyribonucleic acid (DNA) the crucial biomolecule in evolution and metabolism. DNA-like double helical nanostructures are probably the most fantastic yet ubiquitous geometry at the nanoscale level, which are expected to exhibit exceptional and even rather different properties due to the unique organization of the two single helices and their synergistic effect. The organization of nanomaterials into double helical structures is an emerging hot topic for nanomaterials science due to their promising exceptional unique properties and applications. This review focuses on the state-of-the-art research progress for the fabrication of double-helical nanostructures based on `bottom-up' and `top-down' strategies. The relevant nanoscale, mesoscale, and macroscopic scale fabrication methods, as well as the properties of the double helical nanostructures are included. Critical perspectives are devoted to the synthesis principles and potential applications in this emerging research area. A multidisciplinary approach from the scope of nanoscience, physics, chemistry, materials, engineering, and other application areas is still required to the well-controlled and large-scale synthesis, mechanism, property, and application exploration of double helical nanostructures.

  15. Clinical evaluation of BioPlex 2200 HIV Ag-Ab, an automated screening method providing discrete detection of HIV-1 p24 antigen, HIV-1 antibody, and HIV-2 antibody.

    PubMed

    Salmona, Maud; Delarue, Severine; Delaugerre, Constance; Simon, François; Maylin, Sarah

    2014-01-01

    Early and accurate diagnosis is essential for optimal therapeutic outcomes in patients infected with HIV. Currently, none of the commercially available fourth-generation assays differentiate HIV-1 and HIV-2 antibodies (Ab) or the HIV-1 p24 antigen (Ag). The aim of this study was to evaluate the performance of a novel assay, the BioPlex 2200 HIV Ag-Ab. This assay uses a multiplex flow immunoassay design allowing the simultaneous detection and identification of antibodies to HIV-1 (groups M and O), HIV-2, and the HIV-1 p24 antigen, in addition to providing a traditional composite result. A total of 1,505 routine serum samples were prospectively tested. Results were compared with those from the Architect HIV Combo assay. The sensitivity of the BioPlex 2200 was 100%. The specificity assessed on repeated false-positive samples was 99.5%. In addition, 524 frozen specimens from patients known to be infected with HIV-1 or HIV-2 were tested. Of these specimens, 420 were infected with HIV-1, including 156 of known genotypes, 86 were infected with HIV-2, 7 were infected with HIV-1 and HIV-2, and 11 were from patients with acute HIV infection. Sensitivity was 100% for the HIV genotypes tested. The differentiation capabilities of the BioPlex 2200 HIV Ag-Ab assay for HIV-1, HIV-2, dual HIV-1/HIV-2, and early infections were 100%, 90.7%, 100%, and 90.9%, respectively. The BioPlex 2200 is a sensitive and specific assay that offers advantages over conventional HIV combo assays, also referred to as fourth-generation assays, to accurately differentiate and report HIV-1 p24 antigen and HIV-1 and HIV-2 antibodies. PMID:24153130

  16. A novel method based on click chemistry, which overcomes limitations of cell cycle analysis by classical determination of BrdU incorporation, allowing multiplex antibody staining.

    PubMed

    Cappella, Paolo; Gasparri, Fabio; Pulici, Maurizio; Moll, Jürgen

    2008-07-01

    Quantification of BrdU incorporation into DNA is a widely used technique to assess the cell cycle status of cells. DNA denaturation is required for BrdU detection with the drawback that most protein epitopes are destroyed and classical antibody staining techniques for multiplex analysis are not possible. To address this issue we have developed a novel method that overcomes the DNA denaturation step but still allows detection of BrdU. Cells were pulsed for a short time by 5-ethynyl-2'-deoxyuridine, which is incorporated into DNA. The exposed nucleotide alkyne group of DNA was then derivatized in physiologic conditions by the copper (I)-catalyzed azide-alkyne cycloaddition (CuAAC) using BrdU azides. The resulting DNA-bound bromouracil moiety was subsequently detected by commercial anti-BrdU mAb without the need for a denaturation step. Continuous labeling with EdU showed a slightly increased anti-proliferative activity compared to BrdU. However, using a lower concentration of EdU for labeling can compensate for this. Alkynyl tags could be detected quickly by a highly specific reaction using BrdU azides. Fluorescence quenching by the DNA dye PI using both BrdU azides was negligible. Our labeling method is suitable for FCM and HCA and shows a higher signal to noise ratio than other methods. This method also allowed multiplex analysis by simultaneous detection of EdU-BrdU, caspase-3, and phospho-histone 3 mAbs, proving sensitivity and feasibility of this new technique. In addition, it has the potential for use in vivo, as exemplified for bone marrow studies. We have established a new method to determine the position of cells in the cell cycle. This is superior when compared to traditional BrdU detection since it allows multiplex analysis, is more sensitive and shows less quenching with PI. The method provides new opportunities to investigate changes in protein expression at different cell cycle stages using pulse labeling experiments. PMID:18521918

  17. An automated particle counting immunoassay (PACIA) for determination of blocking antibodies against timothy grass pollen in sera from desensitized allergics.

    PubMed

    Djurup, R; Magnusson, C G; Minuva, U; Søndergaard, I; Osterballe, O; Weeke, B

    1983-04-01

    An automated particle counting immunoassay (PACIA) for measurement of blocking antibodies (antigen neutralizing capacity) against timothy grass pollen extract in sera from desensitized allergics is described. Latex particles coated with F(ab')2-anti-timothy are agglutinated by timothy. Serum containing anti-timothy antibodies inhibits the agglutination. Non-agglutinated particles are counted in a modified AutoCounter. Nineteen of 20 sera from timothy allergics who had undergone immunotherapy with purified timothy extract for 30 weeks, showed significant agglutination-inhibition. None of 42 normal human sera gave significant inhibition. The inhibiting antibody could be removed by absorption with protein A and was thus of non-IgE nature, i.e. blocking antibody. The results obtained correlated statistically significantly with those found with a double-antibody method (rS = 0.62, n = 20, t = 3.35, P less than 0.01) and with the cumulated dosage of timothy allergen extract administered to the individual patient (rS = 0.56, n = 20, t = 2.87, P less than 0.02). Between-assay coefficient of variation was from 6.4% to 18.3%. The capacity is 40 samples per hour. The method has also been applied to measurement of blocking antibodies to honey bee and wasp venom. PMID:6846744

  18. Natural monoclonal antibodies and cancer.

    PubMed

    Vollmers, Peter H; Brändlein, Stephanie

    2008-06-01

    Immunity is responsible for recognition and elimination of infectious particles and for removal of cellular waste, modified self structures and transformed cells. Innate or natural immunity acts as a first line defense and is also the link to acquired immunity and memory. By using the human hybridoma technology, a series of monoclonal antibodies and several new tumor-specific targets could be identified. A striking phenomenon of immunity against malignant cells is that all so far isolated tumor-specific antibodies were germ-line coded natural IgM antibodies. And neither in animals nor in humans affinity-maturated tumor-specific IgG antibodies have been detected so far. These IgM's preferentially bind to carbohydrate epitopes on post-transcriptionally modified surface receptors, which are recently patented and preferentially remove malignant cells by inducing apoptosis to avoid inflammatory processes. Our "biology-" or "function-driven" method represents a unique yet powerful approach compared to the typical approaches on screening compounds or antibodies against non-validated targets (mostly differentially expressed). Moreover, the approach creates a competitive patenting strategy of creating proprietary antibodies and validated targets at the same time, which has the potential of further streamlining the discovery of new cancer therapies. PMID:18537750

  19. NEACRP standard problem exercise on criticality codes for dissolving fissile oxides in acids: A reference method for treating the fuel double heterogeneity

    SciTech Connect

    Santamarina, A.; Smith, H.J. . Div. d'Etudes et de Developpement des Reacteurs); Whitesides, G.E. )

    1990-01-01

    The value of international comparison studies by the OECD-NEA Criticality Working Group has again been demonstrated by this study. Computational methods that had been commonly used for criticality safety calculations and which were shown to be valid for systems for which experimental data existed, were demonstrated to be inadequate when extrapolated to some simulated actual situations. The major source of dispersion in the results in international criticality benchmark calculations on problems treating a fuel double heterogeneity is shown to be the incorrect evaluation of effective resonance cross sections for 238U. A reference calculational method is proposed and used to evaluate theoretically the errors created by various standard methods of calculating the effects of self-shielding of resonance cross sections. 7 figs., 3 tabs.

  20. Rapid calculation of diffuse reflectance from a multilayered model by combination of the white Monte Carlo and adding-doubling methods

    PubMed Central

    Yoshida, Kenichiro; Nishidate, Izumi

    2014-01-01

    To rapidly derive a result for diffuse reflectance from a multilayered model that is equivalent to that of a Monte-Carlo simulation (MCS), we propose a combination of a layered white MCS and the adding-doubling method. For slabs with various scattering coefficients assuming a certain anisotropy factor and without absorption, we calculate the transition matrices for light flow with respect to the incident and exit angles. From this series of precalculated transition matrices, we can calculate the transition matrices for the multilayered model with the specific anisotropy factor. The relative errors of the results of this method compared to a conventional MCS were less than 1%. We successfully used this method to estimate the chromophore concentration from the reflectance spectrum of a numerical model of skin and in vivo human skin tissue. PMID:25426319

  1. Isolation of Balamuthia mandrillaris-specific antibody fragments from a bacteriophage antibody display library.

    PubMed

    Siddiqui, Ruqaiyyah; Kulsoom, Huma; Lalani, Salima; Khan, Naveed Ahmed

    2016-07-01

    Balamuthia mandrillaris is a protist pathogen that can cause encephalitis with a mortality rate of more than 95%. Early diagnosis followed by aggressive treatment is a pre-requisite for successful prognosis. Current methods for identifying this organism rely on culture and microscopy, antibody-based methods using animals, or involve the use of molecular tools that are expensive. Here, we describe the isolation of antibody fragments that can be used for the unequivocal identification of B. mandrillaris. B. mandrillaris-specific antibody fragments were isolated from a bacteriophage antibody display library. Individual clones were studied by enzyme-linked immunosorbent assay, and immunofluorescence. Four antibody clones showed specific binding to B. mandrillaris. The usefulness of phage antibody display technology as a diagnostic tool for isolating antibody fragments against B. mandrillaris antigens and studying their biological role(s) is discussed further. PMID:27055361

  2. Antibodies and Selection of Monoclonal Antibodies.

    PubMed

    Hanack, Katja; Messerschmidt, Katrin; Listek, Martin

    2016-01-01

    Monoclonal antibodies are universal binding molecules with a high specificity for their target and are indispensable tools in research, diagnostics and therapy. The biotechnological generation of monoclonal antibodies was enabled by the hybridoma technology published in 1975 by Köhler and Milstein. Today monoclonal antibodies are used in a variety of applications as flow cytometry, magnetic cell sorting, immunoassays or therapeutic approaches. First step of the generation process is the immunization of the organism with appropriate antigen. After a positive immune response the spleen cells are isolated and fused with myeloma cells in order to generate stable, long-living antibody-producing cell lines - hybridoma cells. In the subsequent identification step the culture supernatants of all hybridoma cells are screened weekly for the production of the antibody of interest. Hybridoma cells producing the antibody of interest are cloned by limited dilution till a monoclonal hybridoma is found. This is a very time-consuming and laborious process and therefore different selection strategies were developed since 1975 in order to facilitate the generation of monoclonal antibodies. Apart from common automation of pipetting processes and ELISA testing there are some promising approaches to select the right monoclonal antibody very early in the process to reduce time and effort of the generation. In this chapter different selection strategies for antibody-producing hybridoma cells are presented and analysed regarding to their benefits compared to conventional limited dilution technology. PMID:27236550

  3. Antibody titer

    MedlinePlus

    ... RA, Pincus MR, eds. Henry's Clinical Diagnosis and Management by Laboratory Methods . 22nd ed. Philadelphia, PA: Elsevier ... by: Stuart I. Henochowicz, MD, FACP, Associate Clinical Professor of Medicine, Division of Allergy, Immunology, and Rheumatology, ...

  4. Barberry treatment reduces serum anti-heat shock protein 27 and 60 antibody titres and high-sensitivity c-reactive protein in patients with metabolic syndrome: a double-blind, randomized placebo-controlled trial.

    PubMed

    Zilaee, Marzie; Kermany, Tayyebeh; Tavalaee, Shima; Salehi, Maryam; Ghayour-Mobarhan, Majid; Ferns, Gordon A A

    2014-08-01

    Metabolic syndrome is an important risk factor for cardiovascular disease (CVD). The heat shock proteins (HSPs) are associated with risk factors for CVD. The aim of the present study was to survey the effect of barberry on antibody titres to HSPs and high-sensitivity C-reactive protein (hs-CRP) in patients with metabolic syndrome. In our study, subjects (N=106, 79 women and 27 men, 18-65 years old) with metabolic syndrome were randomized into two groups: a group of patients who received three capsules of barberry and a control group who received three capsules of placebo for 6 weeks. Antibodies against HSPs 27, 60/65 and 70, hs-CRP and lipid profile were determined in patients before (week 0) and after (week 6) intervention. spss software (version 16.0; Inc, Chicago, IL) was used for data analysis. Results showed that barberry had no significant effect on serum level of anti-HSPs 65 and 70. But there was a significant decrease in anti-HSP 27 in both case and control groups (p=0.001 and p<0.001, respectively, in the case and control groups). Barberry decreased significantly anti-HSP 60 in the case group (p=0.03). High-sensitivity CRP was decreased non-significantly (p=0.17) in the case group and increased significantly (p=0.04) in the control group. Barberry decreased significantly low-density lipoprotein and total cholesterol and increased significantly high-density cholesterol (p<0.05). Results of the present study suggested that barberry supplementation in patients with metabolic syndrome decreased significantly anti-HSPs 27 and 60 and hs-CRP levels and improved lipid profile. PMID:24536039

  5. Effect of the double-counting functional on the electronic and magnetic properties of half-metallic magnets using the GGA+U method

    NASA Astrophysics Data System (ADS)

    Tsirogiannis, Christos; Galanakis, Iosif

    2015-11-01

    Methods based on the combination of the usual density functional theory (DFT) codes with the Hubbard models are widely used to investigate the properties of strongly correlated materials. Using first-principle calculations we study the electronic and magnetic properties of 20 half-metallic magnets performing self-consistent GGA+U calculations using both the atomic-limit (AL) and around-mean-field (AMF) functionals for the double counting term, used to subtract the correlation part from the DFT total energy, and compare these results to the usual generalized-gradient-approximation (GGA) calculations. Overall the use of AMF produces results similar to the GGA calculations. On the other hand the effect of AL is diversified depending on the studied material. In general the AL functional produces a stronger tendency towards magnetism leading in some cases to unphysical electronic and magnetic properties. Thus the choice of the adequate double-counting functional is crucial for the results obtained using the GGA+U method.

  6. Quantum dots-based double imaging combined with organic dye imaging to establish an automatic computerized method for cancer Ki67 measurement

    NASA Astrophysics Data System (ADS)

    Wang, Lin-Wei; Qu, Ai-Ping; Liu, Wen-Lou; Chen, Jia-Mei; Yuan, Jing-Ping; Wu, Han; Li, Yan; Liu, Juan

    2016-02-01

    As a widely used proliferative marker, Ki67 has important impacts on cancer prognosis, especially for breast cancer (BC). However, variations in analytical practice make it difficult for pathologists to manually measure Ki67 index. This study is to establish quantum dots (QDs)-based double imaging of nuclear Ki67 as red signal by QDs-655, cytoplasmic cytokeratin (CK) as yellow signal by QDs-585, and organic dye imaging of cell nucleus as blue signal by 4‧,6-diamidino-2-phenylindole (DAPI), and to develop a computer-aided automatic method for Ki67 index measurement. The newly developed automatic computerized Ki67 measurement could efficiently recognize and count Ki67-positive cancer cell nuclei with red signals and cancer cell nuclei with blue signals within cancer cell cytoplasmic with yellow signals. Comparisons of computerized Ki67 index, visual Ki67 index, and marked Ki67 index for 30 patients of 90 images with Ki67 ≤ 10% (low grade), 10% < Ki67 < 50% (moderate grade), and Ki67 ≥ 50% (high grade) showed computerized Ki67 counting is better than visual Ki67 counting, especially for Ki67 low and moderate grades. Based on QDs-based double imaging and organic dye imaging on BC tissues, this study successfully developed an automatic computerized Ki67 counting method to measure Ki67 index.

  7. Quantum dots-based double imaging combined with organic dye imaging to establish an automatic computerized method for cancer Ki67 measurement

    PubMed Central

    Wang, Lin-Wei; Qu, Ai-Ping; Liu, Wen-Lou; Chen, Jia-Mei; Yuan, Jing-Ping; Wu, Han; Li, Yan; Liu, Juan

    2016-01-01

    As a widely used proliferative marker, Ki67 has important impacts on cancer prognosis, especially for breast cancer (BC). However, variations in analytical practice make it difficult for pathologists to manually measure Ki67 index. This study is to establish quantum dots (QDs)-based double imaging of nuclear Ki67 as red signal by QDs-655, cytoplasmic cytokeratin (CK) as yellow signal by QDs-585, and organic dye imaging of cell nucleus as blue signal by 4′,6-diamidino-2-phenylindole (DAPI), and to develop a computer-aided automatic method for Ki67 index measurement. The newly developed automatic computerized Ki67 measurement could efficiently recognize and count Ki67-positive cancer cell nuclei with red signals and cancer cell nuclei with blue signals within cancer cell cytoplasmic with yellow signals. Comparisons of computerized Ki67 index, visual Ki67 index, and marked Ki67 index for 30 patients of 90 images with Ki67 ≤ 10% (low grade), 10% < Ki67 < 50% (moderate grade), and Ki67 ≥ 50% (high grade) showed computerized Ki67 counting is better than visual Ki67 counting, especially for Ki67 low and moderate grades. Based on QDs-based double imaging and organic dye imaging on BC tissues, this study successfully developed an automatic computerized Ki67 counting method to measure Ki67 index. PMID:26839163

  8. The effective encapsulation of a hydrophobic lipid-insoluble drug in solid lipid nanoparticles using a modified double emulsion solvent evaporation method.

    PubMed

    Nabi-Meibodi, Mohsen; Vatanara, Alireza; Najafabadi, Abdolhossein Rouholamini; Rouini, Mohammad Reza; Ramezani, Vahid; Gilani, Kambiz; Etemadzadeh, Seyed Mohammad Hossein; Azadmanesh, Kayhan

    2013-12-01

    Raloxifene HCl (RH), a selective estrogen receptor modulator (SERM), is indicated for the prophylaxis or treatment of postmenopausal osteoporosis. RH shows extremely poor bioavailability due to limited solubility and an extensive intestinal/hepatic first-pass metabolism. Solid lipid nanoparticles (SLNs) are valuable carriers that can enhance drug bioavailability. However, in the case of RH, the encapsulation of the drug in SLNs remains a challenge because of its poor solubility in both water and lipids. In this study, a series of RH-containing SLNs (RH-SLNs) were generated using a modified double emulsion solvent evaporation (DESE) method. Briefly, RH with various drug/lipid ratios was solubilized in the inner core of a double emulsion using different water/organic solvent mixtures. Our best formulation was achieved with the formation of negatively charged nanoparticles, 180nm in diameter, with an encapsulation and loading efficiency of 85% and 4.5%, respectively. It also showed a Fickian mechanism of the drug release in the basic dissolution media. Thermal analysis revealed a distinct decrease in the crystallinity of lipids and RH in comparison with the unprocessed materials. The results of a cell viability assay also showed a better antiproliferative effect of the drug-loaded SLNs versus the free drug solution. Thus, these results indicated that the modified DESE method could be proposed for the effective encapsulation of RH in SLNs with appropriate physicochemical and biological properties. PMID:24036624

  9. Preparation of Nickel-Aluminum-Containing Layered Double Hydroxide Films by Secondary (Seeded) Growth Method and Their Electrochemical Properties.

    PubMed

    Zhang, Fazhi; Guo, Li; Xu, Sailong; Zhang, Rong

    2015-06-23

    Thin films of nickel-aluminum-containing layered double hydroxide (NiAl-LDH) have been prepared on nickel foil and nickel foam substrates by secondary (seeded) growth of NiAl-LDH seed layer. The preparation procedure consists of deposition of LDH seeds from a colloidal suspension on the substrate by dip coating, followed by hydrothermal treatment of the nanocrystals to form the LDH film. The secondary grown film is found to provide a higher crystallinity and more uniform composition of metal cations in the film layer than the in situ grown film on seed-free substrate under identical hydrothermal conditions. A systematic investigation of the film evolution process reveals that the crystallite growth rate is relatively fast for the secondary grown film because of the presence of LDH nanocrystal seeds. Electrochemical performance of the resulting NiAl-LDH films as positive electrode material was further assessed as an example of their practical applications. The secondary grown film electrode delivers improved recharge-discharge capacity and cycling stability compared with that of the in situ grown film, which can be explained by the existence of a unique microstructure of the former. Our findings show an example for the effective fabrication of LDH film with controllable microstructure and enhanced application performance through a secondary (seeded) growth procedure. PMID:26033419

  10. Influence of process parameters on the size distribution of PLA microcapsules prepared by combining membrane emulsification technique and double emulsion-solvent evaporation method.

    PubMed

    Liu, Rong; Ma, Guang-Hui; Wan, Yin-Hua; Su, Zhi-Guo

    2005-11-10

    Relatively uniform-sized biodegradable poly(lactide) (PLA) microcapsules with various sizes were successfully prepared by combining a glass membrane emulsification technique and water-in-oil-in-water (w1/o/w2) double emulsion-solvent evaporation method. A water phase was used as the internal water phase, a mixture solvent of dichloromethane (DCM) and toluene dissolving PLA and Arlacel 83 was used as the oil phase (o). These two solutions were emulsified by a homogenizer to form a w1/o primary emulsion. The primary emulsion was permeated through the uniform pores of a glass membrane into the external water phase by the pressure of nitrogen gas to form the uniform w1/o/w2 double emulsion droplets. Then, the solid polymer microcapsules were obtained by simply evaporating solvent. The influence of process parameters on the size distribution of PLA microcapsules was investigated, with an emphasis on the effect of oil-soluble emulsifier. A unique phenomenon was found that a large part of emulsifier could adsorb on the interface of internal water phase and oil phase, which suppressed its adsorption on the surface of glass membrane, and led to the successful preparation of uniform-sized double emulsion. Finally, by optimizing the process parameters, PLA microcapsules with various sizes having coefficient of variation (CV) value under 14.0% were obtained. Recombinant human insulin (rhI), as a model protein, was encapsulated into the microcapsules with difference sizes, and its encapsulation efficiency and cumulative release were investigated. The result suggested that the release behavior could be simply adjusted just by changing precisely the diameters of microcapsule, benefited from the membrane emulsification technique. PMID:16198091

  11. Double quantization of the regressor space for long-term time series prediction: method and proof of stability.

    PubMed

    Simon, Geoffroy; Lendasse, Amaury; Cottrell, Marie; Fort, Jean-Claude; Verleysen, Michel

    2004-01-01

    The Kohonen self-organization map is usually considered as a classification or clustering tool, with only a few applications in time series prediction. In this paper, a particular time series forecasting method based on Kohonen maps is described. This method has been specifically designed for the prediction of long-term trends. The proof of the stability of the method for long-term forecasting is given, as well as illustrations of the utilization of the method both in the scalar and vectorial cases. PMID:15555859

  12. Magnetic particles functionalized with PAMAM-dendrimers and antibodies: a new system for an ELISA method able to detect Ara h3/4 peanut allergen in foods.

    PubMed

    Speroni, Francesca; Elviri, Lisa; Careri, Maria; Mangia, Alessandro

    2010-08-01

    An innovative enzyme-linked immunosorbent assay (ELISA) format based on antibody-coated magnetic micro-particles (MPs) for the sensitive detection of Ara h3/4 allergen in food is described. The immunosupport is suspended in the incubation solutions and the MPs with the captured allergen can be easily harvested on a magnet, separated from the solutions, and washed using an easy-to-use, fast and selective approach that allows its detection and quantification. Two differently coated MPs, ProteinA-Pn-b and MP-NH(2)-PAMAM G 1.5-Pn-b immunosupports, were tested. The functionalization of the MPs with PAMAM-sodium carboxylate dendrimers elicits a major stability on the immunoglobulin activity resulting in a threefold enhancement of the analytical sensitivity for the assay with respect to a ProteinA immobilization. Validation was carried out on two different matrices: corn flakes and biscuits. In the case of MP-NH(2)-PAMAM G 1.5 -Pn-b immunosupport, limit of detection was found to be 0.2 mg peanuts/kg matrix in both matrices; the linear response range was demonstrated from 2.5 to 15 mg peanuts/kg matrix by performing statistical tests (homoscedasticity and Mandel fitting tests). Good accuracy and recovery (>80 +/- 2%) were obtained. Different food samples were tested and the results were compared with those obtained with a commercially available ELISA kit. The results obtained in this work demonstrated the applicability of the immunomagnetic ELISA methods on real samples and the possibility to perform the assay with significantly reduced reagent and sample consumption. PMID:20607526

  13. FINAL REPORT. ENGINEERED ANTIBODIES FOR MONITORING OF POLYNUCLEAR AROMATIC HYDROCARBONS

    EPA Science Inventory

    This project was conducted to remove the major barrier to the timely development and use of more versatile antibody-based detection and sample cleanup methods. The main objective was to adapt combinatorial antibody library and antibody engineering methods for preparing a panel of...

  14. Mouse monoclonal antibodies against estrogen receptor.

    PubMed

    De Rosa, Caterina; Rossi, Valentina; Abbondanza, Ciro

    2014-01-01

    The production of monoclonal antibodies, by cloning hybridoma derived from the fusion of myeloma cells and spleen lymphocytes, has allowed to obtain great advances in many fields of biological knowledge. The use of specific antibodies to the estrogen receptor, in fact, has been an invaluable method to bring out its mechanisms of action and its effects, both genomic and extra-genomic. Here we describe, step by step, the production of monoclonal antibodies, starting from protocol for antigen preparation to the selection of antibody-secreting hybridoma. PMID:25182770

  15. A rapid method for simultaneous evaluation of free light chain content and aggregate content in culture media of Chinese hamster ovary cells expressing monoclonal antibodies for cell line screening.

    PubMed

    Ishii, Yoichi; Tsukahara, Masayoshi; Wakamatsu, Kaori

    2016-04-01

    The goal of developing a monoclonal antibody (mAb) production process is high productivity and high quality. Because the productivity and quality of mAbs depend on cell line properties, the selection of cell lines suitable for large-scale production is an important stage in process development for mAb production. The light chain (LC) is important for antibody folding and assembly in the endoplasmic reticulum; cell lines that secrete a large amount of LCs in the medium secrete high-quality antibodies with high productivity. LC contents in culture media have been estimated by western blotting, reverse-phase high-performance liquid chromatography, and enzyme-linked immunosorbent assay. However, these analyses require fine tuning of experimental conditions for each antibody analyzed. Here we report a rapid and simple high-sensitivity size-exclusion chromatography (HS-SEC) method to evaluate the contents of low-molecular weight species (LMWS, mainly consisting of LC monomers and dimers) and high-molecular weight species (HMWS, aggregates) in the media for cell line screening. Because LMWS and HMWS are important indicators of productivity and quality, respectively, for cell line screening, HS-SEC will be useful in the first step of cell line selection needed for large-scale production. PMID:26467692

  16. Antibodies and antibody-derived analytical biosensors.

    PubMed

    Sharma, Shikha; Byrne, Hannah; O'Kennedy, Richard J

    2016-06-30

    The rapid diagnosis of many diseases and timely initiation of appropriate treatment are critical determinants that promote optimal clinical outcomes and general public health. Biosensors are now being applied for rapid diagnostics due to their capacity for point-of-care use with minimum need for operator input. Antibody-based biosensors or immunosensors have revolutionized diagnostics for the detection of a plethora of analytes such as disease markers, food and environmental contaminants, biological warfare agents and illicit drugs. Antibodies are ideal biorecognition elements that provide sensors with high specificity and sensitivity. This review describes monoclonal and recombinant antibodies and different immobilization approaches crucial for antibody utilization in biosensors. Examples of applications of a variety of antibody-based sensor formats are also described. PMID:27365031

  17. Antibodies and antibody-derived analytical biosensors

    PubMed Central

    Sharma, Shikha; Byrne, Hannah

    2016-01-01

    The rapid diagnosis of many diseases and timely initiation of appropriate treatment are critical determinants that promote optimal clinical outcomes and general public health. Biosensors are now being applied for rapid diagnostics due to their capacity for point-of-care use with minimum need for operator input. Antibody-based biosensors or immunosensors have revolutionized diagnostics for the detection of a plethora of analytes such as disease markers, food and environmental contaminants, biological warfare agents and illicit drugs. Antibodies are ideal biorecognition elements that provide sensors with high specificity and sensitivity. This review describes monoclonal and recombinant antibodies and different immobilization approaches crucial for antibody utilization in biosensors. Examples of applications of a variety of antibody-based sensor formats are also described. PMID:27365031

  18. Antibody Blood Tests

    MedlinePlus

    ... discovered that people with celiac disease who eat gluten have higher than normal levels of certain antibodies ... rye and barley that are generically known as “gluten.” Antibody Testing: Only A First Step To help ...

  19. RBC Antibody Screen

    MedlinePlus

    ... be limited. Home Visit Global Sites Search Help? RBC Antibody Screen Share this page: Was this page ... Screen Related tests: Direct Antiglobulin Test ; Blood Typing ; RBC Antibody Identification ; Type and Screen; Crossmatch All content ...

  20. Antiparietal cell antibody test

    MedlinePlus

    ... Gastric ulcer - anti-gastric parietal cell antibody; Pernicious anemia - anti-gastric parietal cell antibody; Vitamin B12 - anti- ... may use this test to help diagnose pernicious anemia. Pernicious anemia is a decrease in red blood ...

  1. Lyme disease antibody

    MedlinePlus

    ... JavaScript. The Lyme disease blood test looks for antibodies in the blood to the bacteria that causes ... needed. A laboratory specialist looks for Lyme disease antibodies in the blood sample using the ELISA test . ...

  2. Neonatal Tetanus Immunity in Nigeria: The Effect of HIV Infection on Serum Levels and Transplacental Transfer of Antibodies

    PubMed Central

    Ashir, Mohammed Garba; Rabasa, Adamu Ibrahim; Bukbuk, David Nadeba; Usman, Ahmadu Baba; Mustapha, Modu Gofama; Alhaji, Mohammad Arab

    2016-01-01

    Background. Tetanus toxoid immunisation of pregnant mother has remained the most effective strategy in eliminating neonatal tetanus. Impaired production and/or transplacental transfer of antibodies may affect the effectiveness of this strategy. We studied the effect of maternal HIV infection on serum levels and transplacental transfer of anti-tetanus antibodies. Methods. A total of 162 mother-baby paired serum samples were taken and analysed for anti-tetanus antibody levels using ELISA. Maternal HIV status was also determined by double ELISA technique. Maternal TT vaccination status was also documented. Results. Thirty-eight (23.5%) mothers and 41 (25.3%) babies were seronegative, out of whom 8 mothers were HIV positive and 9 babies were HIV exposed. HIV infected mothers and HIV exposed infants were, respectively, 16.27 times (OR = 16.27, 95% CI = 3.28 to 80.61) and 33.75 times (OR = 33.75, 95% CI = 4.12 to 276.40) more likely to be seronegative for anti-tetanus antibody. Similarly, HIV positive mother-newborn pairs were 7.46 times more likely to have a poor transplacental transfer of tetanus antibodies (OR = 7.46, 95% CI = 1.96 to 28.41). Conclusions. Maternal HIV infection is associated with impaired maternofoetal transfer of anti-tetanus antibodies and seronegativity among mothers and their newborns. Hence, this may hinder efforts to eliminate neonatal tetanus. PMID:26904135

  3. Visualisation of Microglia with the use of Immunohistochemical Double Staining Method for CD-68 and Iba-1 of Cerebral Tissue Samples in Cases of Brain Contusions.

    PubMed

    Stankov, Aleksandar; Belakaposka-Srpanova, Viktorija; Bitoljanu, Natasa; Cakar, Ljupco; Cakar, Zdravko; Rosoklija, Gorazd

    2015-01-01

    In the recent years it has been confirmed that the main component of the immune response in an injury of the nerve cell comes from microglia and macrophages. The main challenge in the field of microglia research is to detect the different stages of cellular activation by visualization of the cell morphology. The existing visualization techniques are based on surface molecules expression in resting and activated microglia cells. For visualization of the microglial cells and their functional state we used double labeling method for cd-68 and iba1 in brain contusions with different survival time. Microglia are stained brown with Iba-1, whereas microglia impregnated with black, grainy color, represents activated microglia stained with CD 68. We had significantly positive results, and we were able to observe changes in the morphology of the microglia that correlated with the survival time. Using double labeling with Iba-1 and cd68 we were able to determine their physiological state based on the morphology and immunoreactivity. PMID:27442380

  4. Double fiber probe with a single fiber Bragg grating based on the capillary-driven self-assembly fabrication method for dimensional measurement of micro parts.

    PubMed

    Cui, Jiwen; Feng, Kunpeng; Hu, Yang; Li, Junying; Dang, Hong; Tan, Jiubin

    2015-12-28

    Focusing on the ultra-precision dimensional measurement of parts with micro-scale dimensions and high aspect ratios, a two-dimensional double fiber probe with a single fiber Bragg grating (DS-FBG probe) is investigated in detail in this paper. The theoretical analysis of the sensing principle is verified by spectrum simulations of the DS-FBG probe with a modified transfer matrix method using the strain distribution within the DS-FBG probe. The fabrication process and physical principle of the capillary-driven self-assembly of double fibers in the UV adhesive with a low viscosity are demonstrated. Experimental results indicate that resolutions of 30 nm in radial direction and 15 nm in axial direction can be achieved, and the short-term displacement drifts within 90 seconds are 28.0 nm in radial direction and 7.9 nm in axial direction, and the long-term displacement drifts within 1 hour are 61.3 nm in radial direction and 17.3 nm in axial direction. The repeatability of the probing system can reach 60 nm and the measurement result of a standard nozzle is 300.49 μm with a standard deviation of 20 nm. PMID:26831960

  5. Rapid photometric detection of thymine residues partially flipped out of double helix as a method for direct scanning of point mutations and apurinic DNA sites.

    PubMed

    Logvina, N A; Yakubovskaya, M G; Dolinnaya, N G

    2011-02-01

    A spectroscopic assay for detection of extrahelical thymine residues in DNA heteroduplexes under their modification by potassium permanganate has been developed. The assay is based on increase in absorbance at 420 nm due to accumulation of thymidine oxidation intermediates and soluble manganese dioxide. The analysis was carried out using a set of 19-bp DNA duplexes containing unpaired thymidines opposite tetrahydrofuranyl derivatives mimicking a widespread DNA damage (apurinic (AP) sites) and a library of 50-bp DNA duplexes containing all types of base mismatches in different surroundings. The relation between the selectivity of unpaired T oxidation and the thermal stability of DNA double helix was investigated. The method described here was shown to discriminate between DNA duplexes with one or two AP sites and to reveal thymine-containing mismatches and all noncanonical base pairs in AT-surroundings. Comparative results of CCM analysis and the rapid photometric assay for mismatch detection are demonstrated for the first time in the same model system. The chemical reactivity of target thymines was shown to correlate with local disturbance of double helix at the mismatch site. As the spectroscopic assay does not require the DNA cleavage reaction and gel electrophoresis, it can be easily automated and used for primary screening of somatic mutations. PMID:21568858

  6. New treatment method for boron in aqueous solutions using Mg-Al layered double hydroxide: Kinetics and equilibrium studies.

    PubMed

    Kameda, Tomohito; Oba, Jumpei; Yoshioka, Toshiaki

    2015-08-15

    Mg-Al layered double hydroxides (LDHs) intercalated with NO3(-) (NO3 · Mg - Al LDHs) and with Cl(-) (Cl · Mg - Al LDHs) were found to take up boron from aqueous solutions. Boron was removed by anion exchange of B(OH)4(-) in solution with NO3(-) and Cl(-) intercalated in the interlayer of the LDH. Using three times the stoichiometric quantity of NO3 · Mg-Al LDH, the residual concentration of B decreased from 100 to 1.9 mg L(-1) in 120 min. Using five times the stoichiometric quantity of Cl · Mg - Al LDH, the residual concentration of B decreased from 100 to 5.6 mg L(-1) in 120 min. It must be emphasized that, in both cases, the residual concentration of B was less than the effluent standards in Japan (10 mg L(-1)). The rate-determining step of B removal by the NO3 · Mg - Al and Cl · Mg - Al LDHs was found to be chemical adsorption involving anion exchange of B(OH)4(-) with intercalated NO3(-) and Cl(-). The removal of B was well described by a pseudo second-order kinetic equation. The adsorption of B by NO3 · Mg - Al LDH and Cl · Mg - Al LDH followed a Langmuir-type adsorption. The values of the maximum adsorption and the equilibrium adsorption constant were 3.6 mmol g(-1) and 1.7, respectively, for NO3 · Mg - Al LDH, and 3.8 mmol g(-1) and 0.7, respectively, for Cl · Mg-Al LDH. The B(OH)4(-) in B(OH)4 · Mg - Al LDH produced by removal of B was found to undergo anion exchange with NO3(-) and Cl(-) in solution. The NO3 · Mg - Al and Cl · Mg - Al LDHs obtained after this regeneration treatment were able to remove B from aqueous solutions, indicating the possibility of recycling NO3 · Mg - Al and Cl · Mg - Al LDHs for B removal. PMID:25827268

  7. Drug delivery system for an anticancer agent, chlorogenate-Zn/Al-layered double hydroxide nanohybrid synthesised using direct co-precipitation and ion exchange methods

    NASA Astrophysics Data System (ADS)

    Barahuie, Farahnaz; Hussein, Mohd Zobir; Arulselvan, Palanisamy; Fakurazi, Sharida; Zainal, Zulkarnain

    2014-09-01

    A nano-structured drug-inorganic clay hybrid involving an active anticancer compound, which is chlorogenic acid (CA) intercalated into Zn/Al-layered double hydroxide, has been assembled via ion-exchange and co-precipitation methods to form a nanohybrid CZAE (a chlorogenic acid-Zn/Al nanohybrid synthesised using an ion-exchange method) and CZAC (a chlorogenic acid-Zn/Al nanohybrid synthesised using a direct method), respectively. The X-ray diffraction (XRD) results confirmed that the CA-LDH had a hybrid structure in which the anionic chlorogenate is arranged between the interlayers as a horizontal monolayer at 90 and 20° angles from the x axis for CZAE and CZAC, respectively. Both nanohybrids have the properties of mesoporous materials. The high loading percentage of chlorogenic acid (approximately 43.2% for CZAE and 45.3% for CZAC) with basal spacings of 11.7 and 12.6 Å for CZAE and CZAC, respectively, corroborates the successful intercalation of chlorogenic acid into the interlayer gallery of layered double hydroxides. Free chlorogenic acid and the synthesised nanocomposites (CZAE, CZAC) were assessed for their cytotoxicity against various cancer cells. The Fourier transform infrared data supported the formation of both nanohybrids, and a thermal analysis showed that the nanohybrids are more thermally stable than their counterparts. The chlorogenate shows a sustained release, and the release rate of chlorogenate from CZAE and CZAC nanohybrids at pH 7.4 is remarkably lower than that at pH 4.8 due to their different release mechanisms. The release rate of chlorogenate from both nanohybrids can be described as pseudo-second order. The present investigation revealed the potential of the nanohybrids to enhance the in vitro anti-tumour effect of chlorogenic acid in liver and lung cancer cells in vitro.

  8. Modeling Antibody Diversity.

    ERIC Educational Resources Information Center

    Baker, William P.; Moore, Cathy Ronstadt

    1998-01-01

    Understanding antibody structure and function is difficult for many students. The rearrangement of constant and variable regions during antibody differentiation can be effectively simulated using a paper model. Describes a hands-on laboratory exercise which allows students to model antibody diversity using readily available resources. (PVD)

  9. Double Layers in Astrophysics

    NASA Technical Reports Server (NTRS)

    Williams, Alton C. (Editor); Moorehead, Tauna W. (Editor)

    1987-01-01

    Topics addressed include: laboratory double layers; ion-acoustic double layers; pumping potential wells; ion phase-space vortices; weak double layers; electric fields and double layers in plasmas; auroral double layers; double layer formation in a plasma; beamed emission from gamma-ray burst source; double layers and extragalactic jets; and electric potential between plasma sheet clouds.

  10. Remembering antibodies coming of age.

    PubMed

    Melchers, Fritz

    2016-01-01

    Fifty years ago, Norbert Hilschmann discovered that antibodies have variable immunoglobulin domains to bind antigens, and constant domains to carry out effector functions in the immune system. Just as this happened, the author of this perspective entered the field of immunology. Ten years later, the genetic basis of antibody variability was discovered by Susumu Tonegawa and his colleagues at the Basel Institute for Immunology, where the author had become a scientific member. At the same time, Georges Köhler, a former graduate student of the author's at the Basel Institute, invented with Cesar Milstein at the Laboratory of Molecular Biology in Cambridge, England, the method to produce monoclonal antibodies. The author describes here his memories connected to these three monumental, paradigm-changing discoveries, which he observed in close proximity. PMID:27144253

  11. Direct visualization method of the atomic structure of light and heavy atoms with double-detector C{sub s}-corrected scanning transmission electron microscopy

    SciTech Connect

    Kotaka, Yasutoshi

    2012-09-24

    The advent of C{sub s}-corrected scanning transmission electron microscopy (STEM) has advanced the observation of atomic structures in materials and nanotechnology devices. High-angle annular dark-field (HAADF)-STEM using an annular detector visualizes heavy elements as bright spots at atomic resolution that can be observed with the Z-contrast technique. In this study, the atomic column of light elements is directly observed as bright spots by middle-angle bright-field (MABF)-STEM imaging. Therefore, a double-detector STEM imaging method was developed, exploiting the advantage of both MABF-STEM and HAADF-STEM to maximum, which consists of multiple exposures of simultaneously observed MABF- and HAADF-STEM images in red-green-blue color.

  12. An efficient formulation and implementation of the analytic energy gradient method to the single and double excitation coupled-cluster wave function - Application to Cl2O2

    NASA Technical Reports Server (NTRS)

    Rendell, Alistair P.; Lee, Timothy J.

    1991-01-01

    The analytic energy gradient for the single and double excitation coupled-cluster (CCSD) wave function has been reformulated and implemented in a new set of programs. The reformulated set of gradient equations have a smaller computational cost than any previously published. The iterative solution of the linear equations and the construction of the effective density matrices are fully vectorized, being based on matrix multiplications. The new method has been used to investigate the Cl2O2 molecule, which has recently been postulated as an important intermediate in the destruction of ozone in the stratosphere. In addition to reporting computational timings, the CCSD equilibrium geometries, harmonic vibrational frequencies, infrared intensities, and relative energetics of three isomers of Cl2O2 are presented.

  13. NDE evaluation of the intergranular corrosion susceptibility of a 2205 duplex stainless steel using thermoelectric power and double loop electrochemical potentiokinetic reactivation methods

    NASA Astrophysics Data System (ADS)

    Ortiz, N.; Carreón, H.; Ruiz, A.

    2013-01-01

    There is a need for a nondestructive technique to assess rapidly and with confidence the degree of sensitization (DOS) in duplex stainless steel (DSS). In this investigation, we present the use of thermoelectric power (TEP) measurements as nondestructive method for the determination of DOS in isothermally aged 2205 DSS at 700°C for different aging times. The DOS of the aged samples was first established by performing the double loop electrochemical potentiokinetic reactivation (DL-EPR) test. The microstructural evolution was evaluated by scanning electron microscopy (SEM). Experimental results indicate that TEP coefficient is sensitive to gradual microstructural changes produced by thermal aging and can be used to monitor IGC sensitization of 2205 duplex stainless steel.

  14. Uses of monoclonal antibody 8H9

    DOEpatents

    Cheung, Nai-Kong V

    2013-08-06

    This invention provides a composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a suitable carrier. This invention provides a pharmaceutical composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a pharmaceutically acceptable carrier. This invention also provides an antibody other than the monoclonal antibody 8H9 comprising the complementary determining regions of monoclonal antibody 8H9 or a derivative thereof, capable of binding to the same antigen as the monoclonal antibody 8H9. This invention provides a substance capable of competitively inhibiting the binding of monoclonal antibody 8H9. This invention also provides an isolated scFv of monoclonal antibody 8H9 or a derivative thereof. This invention also provides the 8H9 antigen. This invention also provides a method of inhibiting the growth of tumor cells comprising contacting said tumor cells with an appropriate amount of monoclonal antibody 8H9 or a derivative thereof.

  15. Uses of monoclonal antibody 8H9

    DOEpatents

    Cheung, Nai-Kong V.

    2010-06-15

    This invention provides a composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a suitable carrier. This invention provides a pharmaceutical composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a pharmaceutically acceptable carrier. This invention also provides an antibody other than the monoclonal antibody 8H9 comprising the complementary determining regions of monoclonal antibody 8H9 or a derivative thereof, capable of binding to the same antigen as the monoclonal antibody 8H9. This invention provides a substance capable of competitively inhibiting the binding of monoclonal antibody 8H9. This invention also provides an isolated scFv of monoclonal antibody 8H9 or a derivative thereof. This invention also provides the 8H9 antigen. This invention also provides a method of inhibiting the growth of tumor cells comprising contacting said tumor cells with an appropriate amount of monoclonal antibody 8H9 or a derivative thereof.

  16. Double screening

    NASA Astrophysics Data System (ADS)

    Gratia, Pierre; Hu, Wayne; Joyce, Austin; Ribeiro, Raquel H.

    2016-06-01

    Attempts to modify gravity in the infrared typically require a screening mechanism to ensure consistency with local tests of gravity. These screening mechanisms fit into three broad classes; we investigate theories which are capable of exhibiting more than one type of screening. Specifically, we focus on a simple model which exhibits both Vainshtein and kinetic screening. We point out that due to the two characteristic length scales in the problem, the type of screening that dominates depends on the mass of the sourcing object, allowing for different phenomenology at different scales. We consider embedding this double screening phenomenology in a broader cosmological scenario and show that the simplest examples that exhibit double screening are radiatively stable.

  17. Reduced Antibody Responses to Vaccinations in Children Exposed to Polychlorinated Biphenyls

    PubMed Central

    Heilmann, Carsten; Grandjean, Philippe; Weihe, Pál; Nielsen, Flemming; Budtz-Jørgensen, Esben

    2006-01-01

    Background Developmental exposure to polychlorinated biphenyls (PCBs) has been implicated as a possible cause of deficient immune function in children. This study was designed to assess whether prenatal and postnatal exposure to PCBs impacts on antibody response to childhood immunizations. Methods and Findings Two birth cohorts were formed in the Faroe Islands, where exposures vary widely, because traditional diets may include whale blubber contaminated with PCBs. Prenatal exposure was determined from maternal concentrations of PCBs in pregnancy serum and milk. Following routine childhood vaccinations against tetanus and diphtheria, 119 children were examined at 18 mo and 129 children at 7 y of age, and their serum samples were analyzed for tetanus and diphtheria toxoid antibodies and for PCBs. The antibody response to diphtheria toxoid decreased at age 18 mo by 24.4% (95% confidence interval [CI], 1.63–41.9; p = 0.04) for each doubling of the cumulative PCB exposure at the time of examination. The diphtheria response was lower at age 7 y and was not associated with the exposure. However, the tetanus toxoid antibody response was affected mainly at age 7 y, decreasing by 16.5% (95% CI, 1.51–29.3; p = 0.03) for each doubling of the prenatal exposure. Structural equation analysis showed that the early postnatal exposure was the most important predictor of a decreased vaccination response. Conclusions Increased perinatal exposure to PCBs may adversely impact on immune responses to childhood vaccinations. The clinical implications of insufficient antibody production emphasize the need for prevention of immunotoxicant exposures. PMID:16942395

  18. Synthesis of layered double hydroxides containing Mg2+, Zn2+, Ca2+ and Al3+ layer cations by co-precipitation methods-A review

    NASA Astrophysics Data System (ADS)

    Theiss, Frederick L.; Ayoko, Godwin A.; Frost, Ray L.

    2016-10-01

    Co-precipitation is a common method for the preparation of layered double hydroxides (LDHs) and related materials. This review article is aimed at providing newcomers to the field with some examples of the types of co-precipitation reactions that have been reported previously and to briefly investigate some of the properties of the products of these reactions. Due to the sheer volume of literature on the subject, the authors have had to limit this article to the synthesis of Mg/Al, Zn/Al and Ca/Al LDHs by co-precipitation and directly related methods. LDHs have been synthesised from various reagents including metal salts, oxides and hydroxides. Co-precipitation is also useful for the direct synthesis of LDHs with a wide range of interlayer anions and various bases have been successfully employed to prepare LDHs. Examples of other synthesis techniques including the urea method, hydrothermal synthesis and various mechanochemical methods that are undoubtedly related to co-precipitation have also been included in this review. The effect of post synthesis hydrothermal has also been summarised.

  19. Teaching the Double Layer.

    ERIC Educational Resources Information Center

    Bockris, J. O'M.

    1983-01-01

    Suggests various methods for teaching the double layer in electrochemistry courses. Topics addressed include measuring change in absolute potential difference (PD) at interphase, conventional electrode potential scale, analyzing absolute PD, metal-metal and overlap electron PDs, accumulation of material at interphase, thermodynamics of electrified…

  20. A novel double recognition enzyme-linked immunosorbent assay based on the nucleocapsid protein for early detection of European porcine reproductive and respiratory syndrome virus infection.

    PubMed

    Venteo, A; Rebollo, B; Sarraseca, J; Rodriguez, M J; Sanz, A

    2012-04-01

    Precise and rapid detection of porcine reproductive respiratory syndrome virus (PRRSV) infection in swine farms is critical. Improvement of control procedures, such as testing incoming gilt and surveillance of seronegative herds requires more rapid and sensitive methods. However, standard serological techniques detect mainly IgG antibodies. A double recognition enzyme-linked immunosorbent assay (DR-ELISA) was developed for detection of antibodies specific to European PRRSV. This new assay can recognize both IgM and IgG antibodies to PRSSV which might be useful for detecting in routine surveillance assays pigs that are in the very early stages of infection and missed by conventional assays detecting only IgG antibodies. DR-ELISA is based on the double recognition of antigen by antibody. In this study, the recombinant nucleocapsid protein (N) of PRRSV was used both as the coating and the enzyme-conjugated antigen. To evaluate the sensitivity of the assay at early stages of the infection, sera from 69 pigs infected with PRRSV were collected during successive days post infection (pi) and tested. While standard methods showed low sensitivity rates before day 14 pi, DR-ELISA detected 88.4% seropositive samples at day 7 showing greater sensitivity at early stages of the infection. Further studies were carried out to assess the efficiency of the new assay, and the results showed DR-ELISA to be a sensitive and accurate method for early diagnosis of EU-PRRSV infection. PMID:22342444

  1. Double arch mirror study

    NASA Technical Reports Server (NTRS)

    Vukobratovich, D.; Hillman, D.

    1983-01-01

    The development of a method of mounting light weight glass mirrors for astronomical telescopes compatible with the goals of the Shuttle Infrared Telescope Facility (SIRTF) was investigated. A 20 in. diameter double arch lightweight mirror previously fabricated was modified to use a new mount configuration. This mount concept was developed and fabricated. The mounting concept of the double mounting mirror is outlined. The modifications made to the mirror, fabrication of the mirror mount, and room temperature testing of the mirror and mount and the extension of the mirror and mount concept to a full size (40 in. diameter) primary mirror for SIRTF are discussed.

  2. A double-spike method for K-Ar measurement: A technique for high precision in situ dating on Mars and other planetary surfaces

    NASA Astrophysics Data System (ADS)

    Farley, K. A.; Hurowitz, J. A.; Asimow, P. D.; Jacobson, N. S.; Cartwright, J. A.

    2013-06-01

    A new method for K-Ar dating using a double isotope dilution technique is proposed and demonstrated. The method is designed to eliminate known difficulties facing in situ dating on planetary surfaces, especially instrument complexity and power availability. It may also have applicability in some terrestrial dating applications. Key to the method is the use of a solid tracer spike enriched in both 39Ar and 41K. When mixed with lithium borate flux in a Knudsen effusion cell, this tracer spike and a sample to be dated can be successfully fused and degassed of Ar at <1000 °C. The evolved 40Ar∗/39Ar ratio can be measured to high precision using noble gas mass spectrometry. After argon measurement the sample melt is heated to a slightly higher temperature (˜1030 °C) to volatilize potassium, and the evolved 39K/41K ratio measured by Knudsen effusion mass spectrometry. Combined with the known composition of the tracer spike, these two ratios define the K-Ar age using a single sample aliquot and without the need for extreme temperature or a mass determination. In principle the method can be implemented using a single mass spectrometer. Experiments indicate that quantitative extraction of argon from a basalt sample occurs at a sufficiently low temperature that potassium loss in this step is unimportant. Similarly, potassium isotope ratios measured in the Knudsen apparatus indicate good sample-spike equilibration and acceptably small isotopic fractionation. When applied to a flood basalt from the Viluy Traps, Siberia, a K-Ar age of 351 ± 19 Ma was obtained, a result within 1% of the independently known age. For practical reasons this measurement was made on two separate mass spectrometers, but a scheme for combining the measurements in a single analytical instrument is described. Because both parent and daughter are determined by isotope dilution, the precision on K-Ar ages obtained by the double isotope dilution method should routinely approach that of a pair of

  3. Antibody profiling sensitivity through increased reporter antibody layering

    DOEpatents

    Apel, William A.; Thompson, Vicki S

    2010-04-13

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  4. Antibody profiling sensitivity through increased reporter antibody layering

    DOEpatents

    Apel, William A.; Thompson, Vicki S.

    2013-02-26

    A method for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method comprises attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to the antigens in the array to form immune complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, to form an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.

  5. Monoclonal antibody specific for a pigmentation associated antigen

    SciTech Connect

    Thomson, T.M.; Mattes, M.J.; Old, L.J.; Lloyd, K.O

    1989-01-17

    Monoclonal antibody TA99, which specifically binds to a pigmentation associated antigen present on melanoma cells is described. Additionally, the hybridoma cell line deposited with the ATCC under Accession Number HB 8704 from which the antibody is derived, as well as methods for using the antibody are described.

  6. A new therapeutic antibody removal method using antigen-positive red cells. II. Application to a P-incompatible pregnant woman.

    PubMed

    Yoshida, H; Ito, K; Emi, N; Kanzaki, H; Matsuura, S

    1984-01-01

    Therapeutic antibody removal using antigen-positive red cells was applied to a pregnant woman who has high titered IgG anti-P and had lost all of 4 previous fetuses in P-incompatible pregnancy. The treatment was commenced at the 6th week of gestation and was intensively performed 93 times up to the 35th week. The volume of plasma treated was 2.3 1 per procedure on the average and totalled 215 1. The antibody titer was kept at a low level. At the end of the 35th week of gestation a female infant was delivered by cesarean section. The baby did not require an exchange transfusion and encountered no complications. PMID:6464420

  7. Study on a Simple Method for Controlling the Engine Output Power of Hybrid Powered Railway Vehicles with Electric Double Layer Capacitors

    NASA Astrophysics Data System (ADS)

    Okano, Shota; Shibuya, Hiroyuki; Kondo, Keiichiro

    This paper presents a simple and energy-saving method for controlling hybrid powered railway vehicles that run on rural non-electrified railway lines and have diesel engine and electrical double layer capacitors (EDLCs). The aim this study is to reduce both the fuel consumption and the capacitance of EDLCs. A basic idea proposed in this paper is that EDLCs supply and absorb the kinetic energy of the vehicle and the engine output compensates supply the energy loss with the vehicle running. Thus, the energy loss is not taken into consideration while expressing the EDLC voltage reference (equation 1); energy loss is considered when the engine is in operating mode. The proposed method is examined by performing numerical simulations for various values of engine operation time, load, and grade section. The results of this study reveal the relationship between the capacitance of the EDLCs and the fuel consumption. Using this proposed control methods, excessive charging of EDLCs can be avoided. The results of this study are expected to expedite the development of energy-saving railway vehicles for the non-electrified lines. Finally, the results of this study increase the possibility of developing hybrid powered railway vehicles.

  8. X-ray Diffraction Measurement of GaInNAs/GaAs Double Quantum Well Structures with Novel Analysis Method for Broadening Factors

    NASA Astrophysics Data System (ADS)

    Nakashima, Kiichi; Tateno, Kouta

    2006-09-01

    The structural deterioration of GaInNAs/GaAs double-quantum-well (DQW) samples was analyzed by X-ray diffraction measurement with a novel peak broadening method. We effectively analyzed broadening properties by taking the difference between 004 X-ray satellite profiles measured with two different types of scanning configuration: a conventional configuration without a receiving slit and that with an analyzer crystal placed in front of a receiving detector. We found that the broadening due to structural deterioration can be simply and clearly analyzed by focusing attention on the difference between two types of profile shape, particularly at the valley minimum parts of satellite patterns. It is demonstrated that the difference at the valley minimum parts clearly represents various aspects of the deterioration of DQW structures. Particularly, it is remarkable that not only the degree of deterioration of the DQW structures but also the change in broadening mechanism due to the proceeding deterioration can be effectively analyzed. We propose and formulate a new simple method for analyzing the difference in valley shape by introducing two characteristic indices representing the difference. It is experimentally demonstrated that the new method is effective in both sensitively detecting and characterizing the deterioration of DQW structures by identifying broadening due to tilt distribution or lateral size effects of mosaic structures. From these results, it is concluded that the analysis of the valley minimum parts of profiles is a simple and effective tool for X-ray diffraction measurement.

  9. Antibody Therapeutics in Oncology

    PubMed Central

    Wold, Erik D; Smider, Vaughn V; Felding, Brunhilde H

    2016-01-01

    One of the newer classes of targeted cancer therapeutics is monoclonal antibodies. Monoclonal antibody therapeutics are a successful and rapidly expanding drug class due to their high specificity, activity, favourable pharmacokinetics, and standardized manufacturing processes. Antibodies are capable of recruiting the immune system to attack cancer cells through complement-dependent cytotoxicity or antibody dependent cellular cytotoxicity. In an ideal scenario the initial tumor cell destruction induced by administration of a therapeutic antibody can result in uptake of tumor associated antigens by antigen-presenting cells, establishing a prolonged memory effect. Mechanisms of direct tumor cell killing by antibodies include antibody recognition of cell surface bound enzymes to neutralize enzyme activity and signaling, or induction of receptor agonist or antagonist activity. Both approaches result in cellular apoptosis. In another and very direct approach, antibodies are used to deliver drugs to target cells and cause cell death. Such antibody drug conjugates (ADCs) direct cytotoxic compounds to tumor cells, after selective binding to cell surface antigens, internalization, and intracellular drug release. Efficacy and safety of ADCs for cancer therapy has recently been greatly advanced based on innovative approaches for site-specific drug conjugation to the antibody structure. This technology enabled rational optimization of function and pharmacokinetics of the resulting conjugates, and is now beginning to yield therapeutics with defined, uniform molecular characteristics, and unprecedented promise to advance cancer treatment. PMID:27081677

  10. Comparison of human immune responses to purified Vero cell and human diploid cell rabies vaccines by using two different antibody titration methods.

    PubMed Central

    Kitala, P M; Lindqvist, K J; Koimett, E; Johnson, B K; Chunge, C N; Perrin, P; Olsvik, O

    1990-01-01

    Antibody responses to a conventional rabies preexposure regimen of a new purified Vero cell rabies vaccine (PVRV) and a human diploid cell vaccine (HDCV) were compared in 80 healthy Kenyan veterinary students. Forty-three of the students received the PVRV and 37 received the HDCV on days 0, 7, and 28. Antibody responses were monitored by using the rapid fluorescent-focus inhibition test (RFFIT) and an inhibition enzyme immunoassay (INH EIA) on days 0, 7, 28, and 49. Both vaccines elicited a rapid antibody response. A good correlation between the RFFIT titers and the INH EIA titers was obtained (r = 0.90). Our results also showed that the INH EIA was more reproducible and might therefore be a suitable substitute for the more expensive and less reproducible RFFIT. The geometric mean titers determined by both tests in the two groups of students were statistically similar during the test period. The RFFIT and the INH EIA gave comparable geometric mean titers, which differed significantly only on day 28 in the PVRV group. The effect of the new PVRV is comparable to that of the more expensive HDCV, as determined by the present test systems. The PVRV could therefore be the vaccine of choice, especially in tropical rabies-endemic areas, where the high cost of the HDCV has confined its use to a privileged few. PMID:2203814

  11. Analysis of anti-neoplastic drug in bacterial ghost matrix, w/o/w double nanoemulsion and w/o nanoemulsion by a validated 'green' liquid chromatographic method.

    PubMed

    Youssof, Abdullah M E; Salem-Bekhit, Mounir M; Shakeel, Faiyaz; Alanazi, Fars K; Haq, Nazrul

    2016-07-01

    The objective of the present investigation was to develop and validate a 'green' reversed phase high-performance liquid chromatography (RP-HPLC) method for rapid analysis of a cytotoxic drug 5-fluorouracil (5-FU) in bulk drug, marketed injection, water-in-oil (w/o) nanoemulsion, double water-in-oil-in-water (w/o/w) nanoemulsion and bacterial ghost (BG) matrix. The chromatography study was carried out at room temperature (25±1°C) using an HPLC system with the help of ultraviolet (UV)-visible detector. The chromatographic performance was achieved with a Nucleodur 150mm×4.6mm RP C8 column filled with 5µm filler as a static phase. The mobile phase consisted of ethyl acetate: methanol (7:3% v/v) which was delivered at a flow rate of 1.0mLmin(-1) and the drug was detected in UV mode at 254nm. The developed method was validated in terms of linearity (r(2)=0.998), accuracy (98.19-102.09%), precision (% RSD=0.58-1.17), robustness (% RSD=0.12-0.53) and sensitivity with satisfactory results. The efficiency of the method was demonstrated by the assay of the drug in marketed injection, w/o nanoemulsion, w/o/w nanoemulsion and BG with satisfactory results. The successful resolution of the drug along with its degradation products clearly established the stability-indicating nature of the proposed method. Overall, these results suggested that the proposed analytical method could be effectively applied to the routine analysis of 5-FU in bulk drug, various pharmaceutical dosage forms and BG. PMID:27154677

  12. Trends in Malignant Glioma Monoclonal Antibody Therapy

    PubMed Central

    Chekhonin, Ivan; Gurina, Olga

    2015-01-01

    Although new passive and active immunotherapy methods are emerging, unconjugated monoclonal antibodies remain the only kind of biological preparations approved for high-grade glioma therapy in clinical practice. In this review, we combine clinical and experimental data discussion. As antiangiogenic therapy is the standard of care for recurrent glioblastoma multiforme (GBM), we analyze major clinical trials and possible therapeutic combinations of bevacizumab, the most common monoclonal antibody to vascular endothelial growth factor (VEGF). Another humanized antibody to gain recognition in GBM is epidermal growth factor (EGFR) antagonist nimotuzumab. Other antigens (VEGF receptor, platelet-derived growth factor receptor, hepatocyte growth factor and c-Met system) showed significance in gliomas and were used to create monoclonal antibodies applied in different malignant tumors. We assess the role of genetic markers (isocitrate dehydrogenase, O6-methylguanine-DNA methyltransnsferase) in GBM treatment outcome prediction. Besides antibodies studied in clinical trials, we focus on perspective targets and briefly list other means of passive immunotherapy.

  13. Complex-scaled equation-of-motion coupled-cluster method with single and double substitutions for autoionizing excited states: Theory, implementation, and examples

    SciTech Connect

    Bravaya, Ksenia B.; Zuev, Dmitry; Epifanovsky, Evgeny; Krylov, Anna I.

    2013-03-28

    Theory and implementation of complex-scaled variant of equation-of-motion coupled-cluster method for excitation energies with single and double substitutions (EOM-EE-CCSD) is presented. The complex-scaling formalism extends the EOM-EE-CCSD model to resonance states, i.e., excited states that are metastable with respect to electron ejection. The method is applied to Feshbach resonances in atomic systems (He, H{sup -}, and Be). The dependence of the results on one-electron basis set is quantified and analyzed. Energy decomposition and wave function analysis reveal that the origin of the dependence is in electron correlation, which is essential for the lifetime of Feshbach resonances. It is found that one-electron basis should be sufficiently flexible to describe radial and angular electron correlation in a balanced fashion and at different values of the scaling parameter, {theta}. Standard basis sets that are optimized for not-complex-scaled calculations ({theta} = 0) are not sufficiently flexible to describe the {theta}-dependence of the wave functions even when heavily augmented by additional sets.

  14. In Situ Determination of Trace Elements in Fish Otoliths by Laser Ablation Double Focusing Sector Field Inductively Coupled Plasma Mass Spectrometry Using a Solution Standard Addition Calibration Method

    NASA Astrophysics Data System (ADS)

    Chen, Z.; Jones, C. M.

    2002-05-01

    Microchemistry of fish otoliths (fish ear bones) is a very useful tool for monitoring aquatic environments and fish migration. However, determination of the elemental composition in fish otolith by ICP-MS has been limited to either analysis of dissolved sample solution or measurement of limited number of trace elements by laser ablation (LA)- ICP-MS due to low sensitivity, lack of available calibration standards, and complexity of polyatomic molecular interference. In this study, a method was developed for in situ determination of trace elements in fish otoliths by laser ablation double focusing sector field ultra high sensitivity Finnigan Element 2 ICP-MS using a solution standard addition calibration method. Due to the lack of matrix-match solid calibration standards, sixteen trace elements (Na, Mg, P, Cr, Mn, Fe, Ni, Cu, Rb, Sr, Y, Cd, La, Ba, Pb and U) were determined using a solution standard calibration with Ca as an internal standard. Flexibility, easy preparation and stable signals are the advantages of using solution calibration standards. In order to resolve polyatomic molecular interferences, medium resolution (M/delta M > 4000) was used for some elements (Na, Mg, P, Cr, Mn, Fe, Ni, and Cu). Both external calibration and standard addition quantification strategies are compared and discussed. Precision, accuracy, and limits of detection are presented.

  15. Novel 5'/3'RACE Method for Amplification and Determination of Single-Stranded RNAs Through Double-Stranded RNA (dsRNA) Intermediates.

    PubMed

    Pankovics, Péter; Boros, Ákos; Reuter, Gábor

    2015-12-01

    To acquire the full-length sequences and to determine the 5'/3'ends of the RNA genomes and mRNA transcripts using the rapid amplification of cDNA ends (RACE) protocols-via cDNA or mRNA templates-are a great challenge. This 4-steps RNA-based RACE method uses different ways to determine the RNA ends through a double-stranded (ds) RNA intermediate (dsRNA-RACE). In the first step a complementary RNA strand is synthesised by Phi6 RNA replicase enzyme next to the template ssRNA forming a dsRNA intermediate. The following steps include adapter ligation, nucleic acid purification and two classical methods with minor modifications reverse transcription and polymerase chain reaction. The dsRNA-RACE protocol could be used in wide variety of ssRNA (cellular, viral, bacterial, etc.) templates in the field of microbiology and cellular biology and suitable for the amplification of full-length RNAs including the 5'/3'ends. This is a novel, expansively utilizable molecular tool with fewer disadvantages than the existing 5'/3'RACE approaches. PMID:26315976

  16. Analytic evaluation of energy gradients for the singles and doubles coupled cluster method including perturbative triple excitations: Theory and applications to FOOF and Cr2

    NASA Astrophysics Data System (ADS)

    Scuseria, Gustavo E.

    1991-01-01

    The analytic energy gradient for the singles and doubles coupled cluster method including a perturbative correction due to triple excitations [CCSD(T)] is formulated and computationally implemented. Encouraged by the recent success in reproducing the experimental equilibrium structure and vibrational frequencies of ozone, the new CCSD(T) gradient method is tested with two other ``difficult'' quantum chemistry problems: FOOF and Cr2. With the largest basis set employed in this work [triple zeta plus two sets of polarization functions (TZ2Pf)] at the CCSD(T) level of theory, the predictions for the O-O and O-F bond lengths in FOOF are 1.218 and 1.589 Å, respectively. These figures are in good agreement with the experimental values 1.216 and 1.575 Å. Based on CCSD calculations with even larger basis sets, it is concluded that the error of 0.014 Å in the O-F bond length at the TZ2Pf/CCSD(T) level of theory is due to the remaining basis set deficiency. On the other hand, the CCSD(T) prediction for the equilibrium bond length of Cr2 (1.604 Å), obtained with a large (10s8p3d2f1g) basis set capable of achieving the Hartree-Fock limit, is still 0.075 Å shorter than experiment, clearly indicating the importance of higher than connected triple excitations in a single-reference treatment of this particular problem.

  17. Simultaneous visualization of extrinsic and intrinsic axon collaterals in Golgi-like detail for mouse corticothalamic and corticocortical cells: a double viral infection method

    PubMed Central

    Watakabe, Akiya; Takaji, Masafumi; Kato, Shigeki; Kobayashi, Kazuto; Mizukami, Hiroaki; Ozawa, Keiya; Ohsawa, Sonoko; Matsui, Ryosuke; Watanabe, Dai; Yamamori, Tetsuo

    2014-01-01

    Here we present a novel tracing technique to stain projection neurons in Golgi-like detail by double viral infection. We used retrograde lentiviral vectors and adeno-associated viral vectors (AAV) to drive “TET-ON/TET-OFF system” in neurons connecting two regions. Using this method, we successfully labeled the corticothalamic (CT) cells of the mouse somatosensory barrel field (S1BF) and motor cortex (M1) in their entirety. We also labeled contra- and ipsilaterally-projecting corticocortical (CC) cells of M1 by targeting contralateral M1 or ipsilateral S1 for retrograde infection. The strength of this method is that we can observe the morphology of specific projection neuron subtypes en masse. We found that the group of CT cells extends their dendrites and intrinsic axons extensively below but not within the thalamorecipient layer in both S1BF and M1, suggesting that the primary target of this cell type is not layer 4. We also found that both ipsi- and contralateral targeting CC cells in M1 commonly exhibit widespread collateral extensions to contralateral M1 (layers 1–6), bilateral S1 and S2 (layers 1, 5 and 6), perirhinal cortex (layers 1, 2/3, 5, and 6), striatum and claustrum. These findings not only strengthened the previous findings of single cell tracings but also extended them by enabling cross-area comparison of CT cells or comparison of CC cells of two different labeling. PMID:25278843

  18. Use of second antibody in radioimmunotherapy

    SciTech Connect

    Begent, R.H.; Bagshawe, K.D.; Pedley, R.B.; Searle, F.; Ledermann, J.A.; Green, A.J.; Keep, P.A.; Chester, K.A.; Glaser, M.G.; Dale, R.G.

    1987-01-01

    In this study, a second antibody was directed against the first antitumor antibody to accelerate clearance of the /sup 131/I-labeled first antibody and improve tumor to normal tissue ratios of radioactivity. The value of this method in improving the therapeutic index of radioimmunotherapy with /sup 131/I-antibody to CEA has been investigated in nude mice bearing xenografts of human colon carcinoma and in 5 patients with colorectal cancer. The xenografts did not become saturated with anti-CEA as the administered dose was increased to therapeutic levels. At these high dose levels, the second antibody increased tumor to blood ratios to a maximum of 155:1, 48 times the level in controls that did not receive the second antibody. In 5 patients given 50 mCi of anti-CEA, there was no significant toxicity with the second antibody; clearance of radioactivity was accelerated; and tumor imaging was enhanced. The second antibody appears to have the potential to improve the therapeutic index of radioimmunotherapy.

  19. Design friendly double patterning

    NASA Astrophysics Data System (ADS)

    Yesilada, Emek

    2012-03-01

    Double patterning using 193nm immersion has been adapted as the solution to enable 2x nm technology nodes until the arrival of EUV tools. As a result the past few years have seen a huge effort in creating double patterning friendly design flows. These flows have so far proposed a combination of decomposition rules at cell level and/or at placement level as well as sophisticated decomposition tools with varying density, design iteration and decomposition complexity penalties. What is more, designers have to familiarize themselves with double patterning challenges and decomposition tools. In this paper an alternative approach is presented that allows the development of dense standard cells with minimal impact on design flow due to double patterning. A real case study is done on 20nm node first metal layer where standard cells are designed without considering decomposition restrictions. The resulting layout is carefully studied in order to establish decomposition or color rules that can map the layout into two masks required for double patterning but without the need of complex coloring algorithms. Since the rules are derived from a decomposition unaware design they do not in return impose heavy restrictions on the design at the cell or placement level and show substantial density gains compared to previously proposed methods. Other key advantages are a simplified design flow without complex decomposition tools that can generate a faster time to market solution all at the same time keeping designers isolated from the challenges of the double patterning. The derived design rules highlight process development path required for design driven manufacturing.

  20. Modified hemi-double-stapling technique combined with the temporal abdominal-wall-lift method for performing Billroth I anastomosis after laparoscopically assisted distal gastrectomy.

    PubMed

    Fujii, Hidenori; Aotake, Toshiharu; Kawakami, Yoshiyuki; Okuda, Yukihiro; Doi, Koji; Hirose, Yuki; Matsushita, Toshio

    2008-12-01

    The authors have used a modified hemi-double-stapling (HDS) technique for reconstruction after laparoscopically assisted distal gastrectomy. The stomach is resected from the greater curvature side using a linear stapler inserted into the stomach from that side at a position vertical to the line of the greater curvature. Resection of the stomach is performed by extending the resection line to the lesser curvature using laparoscopic coagulating shears. The resected specimen is examined. After placement of a purse-string suture at the duodenal stump, an anvil is inserted into the stump, and an additional suture with 2-0 silk is placed over the purse-string suture. A curved intraluminal stapler (CDH25) is inserted into the stomach through the opening made on the lesser curvature side, and the center rod of the stapler is passed through the gastric wall on the corner of the resection line at the greater curvature. Ligation with 2-0 silk is added to the center rod by suturing the gastric tissue 5-8 mm from the center rod to encircle it. The authors call this the "one-knot setup HDS," and with this method, a large-caliber anastomosis is secured. In many cases, it is difficult to observe the anastomotic site through the small incisional opening. However, under laparoscopy with the temporal abdominal wall-lift method using the Multi Flap Gate, the anastomotic site can be easily and safely observed. One-knot setup HDS combined with the temporal abdominal wall-lift method is considered a safe and simple method for performing Billroth I anastomosis in laparoscopic distal gastrectomy. PMID:18324439