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Sample records for antibody identifies tn

  1. Comblike dendrimers containing Tn antigen modulate natural killing and induce the production of Tn specific antibodies

    Czech Academy of Sciences Publication Activity Database

    Vepřek, Pavel; Hajdúch, M.; Džubák, P.; Kulík, R.; Poláková, J.; Bezouška, Karel

    2006-01-01

    Roč. 49, č. 21 (2006), s. 6400-6407 ISSN 0022-2623 R&D Projects: GA AV ČR IAA4020213; GA AV ČR IAA5020403; GA ČR GA304/06/1691 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50200510 Keywords : Tn anti gen * glycodendrimer * NKP-P1 Subject RIV: CE - Biochemistry Impact factor: 5.115, year: 2006

  2. A Tn5051-like mer-containing transposon identified in a heavy metal tolerant strain Achromobacter sp. AO22

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    Bhave Mrinal

    2009-03-01

    Full Text Available Abstract Background Achromobacter sp. AO22 (formerly Alcaligenes sp. AO22, a bacterial strain isolated from a lead-contaminated industrial site in Australia, was previously found to be resistant to moderate to high levels of mercury, copper and other heavy metals. However, the nature and location of the genetic basis for mercuric ion resistance in this strain, had not been previously identified. Findings Achromobacter sp. AO22 contains a functional mer operon with all four essential genes (merRTPA and shows >99% DNA sequence identity to that of Tn501. The mer operon was present on a transposon, designated TnAO22, captured by introducing a broad-host-range IncP plasmid into Achromobacter sp. AO22 and subsequently transferring it to E. coli recipients. The transposition frequency of TnAO22 was 10-2 to 10-3 per target plasmid transferred. Analysis of TnAO22 sequence revealed it belonged to the Tn21 subgroup of the Tn3 superfamily of transposons, with the transposition module having >99% identity with Tn5051 of a Pseudomonas putida strain isolated from a water sample in New York. Conclusion TnAO22 is thus a new variant of Tn5051 of the Tn3 superfamily and the transposon and its associated mercury resistance system are among the few such systems reported in a soil bacterium. Achromobacter sp. AO22 can thus be exploited for applications such as in situ mercury bioremediation of contaminated sites, or the mobile unit and mer operon could be mobilized to other bacteria for similar purposes.

  3. Elaboration and quality control of the inoculum of the experimental vaccine Brucella S19-tn7-GFP for use in white animals and associated serological test for the detection of anti-GFP antibodies

    International Nuclear Information System (INIS)

    Salas Alfaro, Dariana

    2014-01-01

    The preparation of the inoculum of the experimental vaccine Brucella S19-Tn7-GFP is optimized for application in white animals. An associated serological test has allowed differentiating infected animals from those vaccinated with the experimental strain. The same bacteriological and biological properties of the B. abortus S19-Tn7-GFP strain have maintained in the parental vaccine strain S19 and is stable over time. A protocol for the inoculums of strain S19-Tn7-GFP is established for its preparation and use in white animals and quality control. The inoculum stability is evaluated through the simulation of conditions that can be presented in the transportation and application process in the field. An enzyme immunoassay ELISA is optimized for the detection of anti-GFP antibodies in cattle [es

  4. The minimal replicon of the Streptomyces ghanaensis plasmid pSG5 identified by subcloning and Tn5 mutagenesis.

    Science.gov (United States)

    Muth, G; Wohlleben, W; Pühler, A

    1988-03-01

    The cryptic plasmid pSG5 of Streptomyces ghanaensis 5/1B (DSM 2932) was characterized to have a molecular size of 12.7 kb and approximate copy number of 20-50 per chromosome. A bifunctional derivative, designated pSW344E, consisting of pSG5 and an Escherichia coli vector plasmid was constructed. Following Tn5 mutagenesis in E. coli, the replication functions of the mutagenized pSW344E plasmids were analysed in S.lividans. A 2 kb DNA fragment of the pSG5 replicon was found to carry replication functions. Subcloning of pSG5 DNA into various replication probe vectors resulted in the identification of the pSG5 minimal replicon, identical to the above mentioned 2 kb DNA region. Several small bifunctional plasmids, able to replicate in E. coli as well as in Streptomyces, were generated during subcloning. Some of these plasmids were found to be useful shuttle vectors.

  5. Analysis of Tn antigenicity with a panel of new IgM and IgG1 monoclonal antibodies raised against leukemic cells

    DEFF Research Database (Denmark)

    Blixt, Klas Ola; Lavrova, Olga I; Mazurov, Dmitriy V

    2012-01-01

    NAc residue of the Tn antigen irrespective of the peptide context or with low selectivity to the glycoproteins. In contrast, IgG mAbs recognized the Tn antigen in the context of a specific peptide motif. Particularly, JA3 mAb reacted to the GSPP or GSPAPP, and JA5 mAb recognized specifically the GSP motif...

  6. Using Tn-seq To Identify Pigmentation-Related Genes of Porphyromonas gingivalis: Characterization of the Role of a Putative Glycosyltransferase.

    Science.gov (United States)

    Klein, Brian A; Cornacchione, Louis P; Collins, Marisha; Malamy, Michael H; Duncan, Margaret J; Hu, Linden T

    2017-07-15

    Cellular pigmentation is an important virulence factor of the oral pathogen Porphyromonas gingivalis Pigmentation has been associated with many bacterial functions, including but not limited to colonization, maintaining a local anaerobic environment by binding oxygen molecules, and defense against reactive oxygen species (ROS) produced by immune cells. Pigmentation-associated loci identified to date have involved lipopolysaccharide, fimbriae, and heme acquisition and processing. We utilized a transposon mutant library of P. gingivalis strain ATCC 33277 and screened for pigmentation-defective colonies using massively parallel sequencing of the transposon junctions (Tn-seq) to identify genes involved in pigmentation. Transposon insertions at 235 separate sites, located in 67 genes and 15 intergenic regions, resulted in altered pigmentation: 7 of the genes had previously been shown to be involved in pigmentation, while 75 genes and intergenic regions had not. To further confirm identification, we generated a smaller transposon mutant library in P. gingivalis strain W83 and identified pigment mutations in several of the same loci as those identified in the screen in ATCC 33277 but also eight that were not identified in the ATCC 33277 screen. PGN_0361/PG_0264, a putative glycosyltransferase gene located between two tRNA synthetase genes and adjacent to a miniature inverted-repeat transposable element, was identified in the Tn-seq screen and then verified through targeted deletion and complementation. Deletion mutations in PGN_0361/PG_0264 glycosyltransferase abolish pigmentation, modulate gingipain protease activity, and alter lipopolysaccharide. The mechanisms of involvement in pigmentation for other loci identified in this study remain to be determined, but our screen provides the most complete survey of genes involved in pigmentation to date. IMPORTANCE P. gingivalis has been implicated in the onset and progression of periodontal disease. One important virulence

  7. Simple mucin-type Tn and sialosyl-Tn carbohydrate antigens in salivary gland carcinomas

    DEFF Research Database (Denmark)

    Therkildsen, M H; Mandel, U; Christensen, M

    1993-01-01

    : Mucin-type Tn and sialosyl-Tn may be regarded as markers of a glandular differentiation pattern in salivary gland carcinomas. The cellular location of the antigen-antibody complex indicates that they are synthesized and secreted from the tumor cells into saliva or serum.......-Tn, which are normally cryptic in human cells and secretions, including saliva and salivary glands. METHODS: Paraffin sections from 50 salivary gland carcinomas of different histologic types were investigated with immunohistologic studies and a panel of monoclonal antibodies with well-defined specificity...

  8. MONOCLONAL ANTIBODIES TO IDENTIFY TOMATO MOSAIC TOBAMOVIRUS (TOMV

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    Duarte Keila M.R.

    2001-01-01

    Full Text Available Monoclonal antibodies were obtained against Tomato mosaic tobamovirus (ToMV isolated in Brazil. One antibody (8G7G2 isotyped as IgG2b (kappa light chain showed strong specificity and very low cross reaction with the Tobacco mosaic virus (TMV. It can be used in identification of tomato mosaic virus (ToMV.

  9. A stable reagent system for screening and identifying red blood cell irregular antibodies: application to commercial antibodies.

    Science.gov (United States)

    Million, L; Pellerin, C; Marchand-Arvier, M; Vigneron, C

    1998-01-01

    Development of a new solid-phase system for screening and identifying irregular red cell antibodies. Red blood cell membranes were prepared by a semi-automated procedure in which the hemolysate solution was passed through a hollow-fiber system. The membranes were fixed to the solid phase (microtiter plates) by centrifugation and incubated with 8% fat-free milk. Antibodies added to the microtiter plate were detected by anti-human antibodies adsorbed onto yellow latex particles. The system had good sensitivity (titer antibodies that are important in transfusion.

  10. Simple mucin-type Tn and sialosyl-Tn carbohydrate antigens in salivary gland carcinomas

    DEFF Research Database (Denmark)

    Therkildsen, M H; Mandel, U; Christensen, M

    1993-01-01

    -Tn, which are normally cryptic in human cells and secretions, including saliva and salivary glands. METHODS: Paraffin sections from 50 salivary gland carcinomas of different histologic types were investigated with immunohistologic studies and a panel of monoclonal antibodies with well-defined specificity......: Mucin-type Tn and sialosyl-Tn may be regarded as markers of a glandular differentiation pattern in salivary gland carcinomas. The cellular location of the antigen-antibody complex indicates that they are synthesized and secreted from the tumor cells into saliva or serum.......BACKGROUND: Neoplastic transformation is associated frequently with changes in the glycosylation process. Simple mucin-type glycosylation in cancer cells has been found to be characterized by incomplete synthesis with precursor accumulation, leading to the exposure of the structures Tn and sialosyl...

  11. Simple mucins (T, sialosyl-T, Tn and sialosyl-Tn) are not diagnostic for malignant breast lesions

    DEFF Research Database (Denmark)

    Reed, W; Bryne, M; Clausen, H

    1994-01-01

    Immunohistochemical study of the distribution of carbohydrate core-structures on O-linked glycoproteins (T, sialosyl-T, Tn and sialosyl-Tn) was performed using specific monoclonal antibodies on 148 primary breast lesions, including 10 normal breast tissues, 16 benign lesions and 122 invasive carc...

  12. Chemoenzymatically synthesized multimeric Tn/STn MUC1 glycopeptides elicit cancer-specific anti-MUC1 antibody responses and override tolerance

    DEFF Research Database (Denmark)

    Sørensen, Anne Louise; Reis, Celso A; Tarp, Mads A

    2005-01-01

    The MUC1 mucin represents a prime target antigen for cancer immunotherapy because it is abundantly expressed and aberrantly glycosylated in carcinomas. Attempts to generate strong humoral immunity to MUC1 by immunization with peptides have generally failed partly because of tolerance. In this stu...... immune response was generated and shown to have the same remarkable cancer specificity. This antibody may hold promise in diagnostic and immunopreventive measures.......The MUC1 mucin represents a prime target antigen for cancer immunotherapy because it is abundantly expressed and aberrantly glycosylated in carcinomas. Attempts to generate strong humoral immunity to MUC1 by immunization with peptides have generally failed partly because of tolerance. In this study...... an optimal vaccine design. Glycopeptides with complete O-glycan occupancy (five sites per repeat) elicited the strongest antibody response reacting with MUC1 expressed in breast cancer cell lines in both Balb/c and MUC1.Tg mice. The elicited humoral immune response showed remarkable specificity for cancer...

  13. Novel exons and splice variants in the human antibody heavy chain identified by single cell and single molecule sequencing.

    Directory of Open Access Journals (Sweden)

    Christopher Vollmers

    Full Text Available Antibody heavy chains contain a variable and a constant region. The constant region of the antibody heavy chain is encoded by multiple groups of exons which define the isotype and therefore many functional characteristics of the antibody. We performed both single B cell RNAseq and long read single molecule sequencing of antibody heavy chain transcripts and were able to identify novel exons for IGHA1 and IGHA2 as well as novel isoforms for IGHM antibody heavy chain.

  14. Fraction A of armadillo submandibular glycoprotein and its desialylated product as sialyl-Tn and Tn receptors for lectins.

    Science.gov (United States)

    Wu, A M; Shen, F; Herp, A; Song, S C; Wu, J H

    1995-02-27

    Fraction A of the armadillo submandibular glycoprotein (ASG-A) is one of the simplest glycoproteins among mammalian salivary mucins. The carbohydrate side chains of this mucous glycoprotein have one-third of the NeuAc alpha 2-->6GalNAc (sialyl-Tn) sequence and two thirds of Tn (GalNAc alpha-->Ser/Thr) residues. Those of the desialylated product (ASG-Tn) are almost exclusively unsubstituted GalNAc residues (Tn determinant). When the binding properties of these glycoproteins were tested by a precipitin assay with Gal, GalNAc and GlcNAc specific lectins, it was found that ASG-Tn reacted strongly with all of the Tn-active lectins and completely precipitated Vicia villosa (VVL both B4 and mixture of A and B), Maclura pomifera (MPA), and Artocarpus integrifolia (jacalin) lectins. However, it precipitated poorly or negligibly with Ricinus communis (RCA1); Dolichos biflorus (DBA); Viscum album, ML-I; Arachis hypogaea (PNA), and Triticum vulgaris (WGA). The reactivity of ASG-A (sialyl-Tn) was as active as that of ASG-Tn with MPA and less or slightly less active than that of ASG-Tn with VVL-A+B, VVL-B4, HPA, WFA, and jacalin, as one-third of its Tn was sialylated. These findings indicate that ASG-A and its desialylated product (ASG-Tn) are highly useful reagents for the differentiation of Tn, T (Gal beta 1-->3GalNAc), A (GalNAc alpha 1-->3Gal) or Gal specific lectins and monoclonal antibodies against such epitopes.

  15. High-content Analysis of Antibody Phage-display Library Selection Outputs Identifies Tumor Selective Macropinocytosis-dependent Rapidly Internalizing Antibodies*

    Science.gov (United States)

    Ha, Kevin D.; Bidlingmaier, Scott M.; Zhang, Yafeng; Su, Yang; Liu, Bin

    2014-01-01

    Many forms of antibody-based targeted therapeutics, including antibody drug conjugates, utilize the internalizing function of the targeting antibody to gain intracellular entry into tumor cells. Ideal antibodies for developing such therapeutics should be capable of both tumor-selective binding and efficient endocytosis. The macropinocytosis pathway is capable of both rapid and bulk endocytosis, and recent studies have demonstrated that it is selectively up-regulated by cancer cells. We hypothesize that receptor-dependent macropinocytosis can be achieved using tumor-targeting antibodies that internalize via the macropinocytosis pathway, improving potency and selectivity of the antibody-based targeted therapeutic. Although phage antibody display libraries have been utilized to find antibodies that bind and internalize to target cells, no methods have been described to screen for antibodies that internalize specifically via macropinocytosis. We hereby describe a novel screening strategy to identify phage antibodies that bind and rapidly enter tumor cells via macropinocytosis. We utilized an automated microscopic imaging-based, High Content Analysis platform to identify novel internalizing phage antibodies that colocalize with macropinocytic markers from antibody libraries that we have generated previously by laser capture microdissection-based selection, which are enriched for internalizing antibodies binding to tumor cells in situ residing in their tissue microenvironment (Ruan, W., Sassoon, A., An, F., Simko, J. P., and Liu, B. (2006) Identification of clinically significant tumor antigens by selecting phage antibody library on tumor cells in situ using laser capture microdissection. Mol. Cell. Proteomics. 5, 2364–2373). Full-length human IgG molecules derived from macropinocytosing phage antibodies retained the ability to internalize via macropinocytosis, validating our screening strategy. The target antigen for a cross-species binding antibody with a highly

  16. High-content analysis of antibody phage-display library selection outputs identifies tumor selective macropinocytosis-dependent rapidly internalizing antibodies.

    Science.gov (United States)

    Ha, Kevin D; Bidlingmaier, Scott M; Zhang, Yafeng; Su, Yang; Liu, Bin

    2014-12-01

    Many forms of antibody-based targeted therapeutics, including antibody drug conjugates, utilize the internalizing function of the targeting antibody to gain intracellular entry into tumor cells. Ideal antibodies for developing such therapeutics should be capable of both tumor-selective binding and efficient endocytosis. The macropinocytosis pathway is capable of both rapid and bulk endocytosis, and recent studies have demonstrated that it is selectively up-regulated by cancer cells. We hypothesize that receptor-dependent macropinocytosis can be achieved using tumor-targeting antibodies that internalize via the macropinocytosis pathway, improving potency and selectivity of the antibody-based targeted therapeutic. Although phage antibody display libraries have been utilized to find antibodies that bind and internalize to target cells, no methods have been described to screen for antibodies that internalize specifically via macropinocytosis. We hereby describe a novel screening strategy to identify phage antibodies that bind and rapidly enter tumor cells via macropinocytosis. We utilized an automated microscopic imaging-based, High Content Analysis platform to identify novel internalizing phage antibodies that colocalize with macropinocytic markers from antibody libraries that we have generated previously by laser capture microdissection-based selection, which are enriched for internalizing antibodies binding to tumor cells in situ residing in their tissue microenvironment (Ruan, W., Sassoon, A., An, F., Simko, J. P., and Liu, B. (2006) Identification of clinically significant tumor antigens by selecting phage antibody library on tumor cells in situ using laser capture microdissection. Mol. Cell. Proteomics. 5, 2364-2373). Full-length human IgG molecules derived from macropinocytosing phage antibodies retained the ability to internalize via macropinocytosis, validating our screening strategy. The target antigen for a cross-species binding antibody with a highly active

  17. Computational analysis of anti-HIV-1 antibody neutralization panel data to identify potential functional epitope residues.

    Science.gov (United States)

    West, Anthony P; Scharf, Louise; Horwitz, Joshua; Klein, Florian; Nussenzweig, Michel C; Bjorkman, Pamela J

    2013-06-25

    Advances in single-cell antibody cloning methods have led to the identification of a variety of broadly neutralizing anti-HIV-1 antibodies. We developed a computational tool (Antibody Database) to help identify critical residues on the HIV-1 envelope protein whose natural variation affects antibody activity. Our simplifying assumption was that, for a given antibody, a significant portion of the dispersion of neutralization activity across a panel of HIV-1 strains is due to the amino acid identity or glycosylation state at a small number of specific sites, each acting independently. A model of an antibody's neutralization IC50 was developed in which each site contributes a term to the logarithm of the modeled IC50. The analysis program attempts to determine the set of rules that minimizes the sum of the residuals between observed and modeled IC50 values. The predictive quality of the identified rules may be assessed in part by whether there is support for rules within individual viral clades. As a test case, we analyzed antibody 8ANC195, an anti-glycoprotein gp120 antibody of unknown specificity. The model for this antibody indicated that several glycosylation sites were critical for neutralization. We evaluated this prediction by measuring neutralization potencies of 8ANC195 against HIV-1 in vitro and in an antibody therapy experiment in humanized mice. These experiments confirmed that 8ANC195 represents a distinct class of glycan-dependent anti-HIV-1 antibody and validated the utility of computational analysis of neutralization panel data.

  18. Identifying the Conditions Under Which Antibodies Protect Against Infection by Equine Infectious Anemia Virus

    OpenAIRE

    Schwartz, Elissa J.; Smith?, Robert J.

    2014-01-01

    The ability to predict the conditions under which antibodies protect against viral infection would transform our approach to vaccine development. A more complete understanding is needed of antibody protection against lentivirus infection, as well as the role of mutation in resistance to an antibody vaccine. Recently, an example of antibody-mediated vaccine protection has been shown via passive transfer of neutralizing antibodies before equine infectious anemia virus (EIAV) infection of horses...

  19. Interaction of hamster submaxillary sialyl-Tn and Tn glycoproteins with Gal, GalNAc and GlcNAc specific lectins.

    Science.gov (United States)

    Wu, A M; Shen, F; Herp, A; Wu, J H

    1994-04-01

    Hamster submaxillary glycoprotein (HSM), one of the simplest glycoproteins among mammalian salivary mucins, is composed of approximately equivalent amounts of protein, hexosamine and sialic acid. The Thr and Ser residues in the protein core account for more than half of all of the amino acid residues, while Lys, Glu, Pro and Ala are the major components of the remaining portion of amino acids. The carbohydrate side chains of this mucous glycoprotein have mainly the NeuAc-GalNAc-(sialyl-Tn) sequence (HSM), and those of the desialylated product (HSM-Tn) are almost exclusively unsubstituted GalNAc residues (Tn determinants). The binding properties of sialyl-Tn (HSM) and asialo-HSM (HSM-Tn) glycoproteins were tested by precipitin assay with Gal, GalNAc and GlcNAc specific lectins. The HSM-Tn completely precipitated Vicia villosa (VVL both B4 and mixture of A and B), Maclura pomifera (MPL), and Artocarpus integrifolia (Jacalin) lectins; less than 2 micrograms of HSM-Tn were required for precipitating 50% of 5.0-6.3 micrograms lectin nitrogen added. HSM-Tn also reacted well with Helix pomatia lectin (HPL), Wistaria floribunda lectin (WFL) and Abrus precatorius agglutinin (APA) and precipitated in each case over 81% of the lectin nitrogen added. The reactivity of HSM-Tn with other lectins (Ricinus communis, RCA1; Dolichol biflorus, DBL; Viscum album, ML-I; Arachis hypogaea, PNA, and Triticum vulgaris, WGA) was weak or negligible. The activity of sialyl-Tn (HSM) was more restricted; HSM reacted well with Jacalin, moderately with MPL and VVL-B4, but was inactive or only weakly with the other lectins used. These findings indicate that HSM and its desialylated product (HSM-Tn) are highly useful reagents for the differentiation of Tn and T/Gal specific lectins and for anti-T, Tn and Af monoclonal antibodies.

  20. Optimizing the Measurement of Colostrum Antibody Concentrations for Identifying BVDV Persistently Infected Calves.

    Science.gov (United States)

    Jenvey, Caitlin J; Reichel, Michael P; Lanyon, Sasha R; Cockcroft, Peter D

    2015-03-09

    Colostrum contains substantially higher concentrations of immunoglobulins compared to serum, which may help to improve the utility of diagnostic tests. The aim of this study was to determine the diagnostic value of colostrum antibody concentrations in identifying Bovine Viral Diarrhoea Virus (BVDV) PI (persistently infected) calf carrying beef heifers following an experimental infection. Colostrum was collected within 12 hours of parturition and tested in undiluted, 1:5, 1:10, 1:100, 1:200, and 1:500 dilutions using an enzyme-linked immunosorbent assay (ELISA) for BVDV antibody. Cows were determined to be carrying a PI calf based on positive quantitative Real Time-Polymerase Chain Reaction and antigen ELISA result on pre-colostral serum and ear notch samples collected from their calf. The median ELISA sample-to-positive (S/P) ratio for colostrum collected from heifers that carried a PI calf were significantly higher than the median ELISA S/P ratio for colostrum collected from heifers that did not carry a PI calf at dilutions of 1:100, 1:200, and 1:500. This study provides further evidence for increased antigenic stimulation in utero by the BVDV viraemic PI calf, which can also be identified with 100% diagnostic sensitivity when using 1:500 dilution colostrum.

  1. Optimizing the Measurement of Colostrum Antibody Concentrations for Identifying BVDV Persistently Infected Calves

    Directory of Open Access Journals (Sweden)

    Caitlin J. Jenvey

    2015-03-01

    Full Text Available Colostrum contains substantially higher concentrations of immunoglobulins compared to serum, which may help to improve the utility of diagnostic tests. The aim of this study was to determine the diagnostic value of colostrum antibody concentrations in identifying Bovine Viral Diarrhoea Virus (BVDV PI (persistently infected calf carrying beef heifers following an experimental infection. Colostrum was collected within 12 hours of parturition and tested in undiluted, 1:5, 1:10, 1:100, 1:200, and 1:500 dilutions using an enzyme-linked immunosorbent assay (ELISA for BVDV antibody. Cows were determined to be carrying a PI calf based on positive quantitative Real Time-Polymerase Chain Reaction and antigen ELISA result on pre-colostral serum and ear notch samples collected from their calf. The median ELISA sample-to-positive (S/P ratio for colostrum collected from heifers that carried a PI calf were significantly higher than the median ELISA S/P ratio for colostrum collected from heifers that did not carry a PI calf at dilutions of 1:100, 1:200, and 1:500. This study provides further evidence for increased antigenic stimulation in utero by the BVDV viraemic PI calf, which can also be identified with 100% diagnostic sensitivity when using 1:500 dilution colostrum.

  2. Chemical Screens Identify Drugs that Enhance or Mitigate Cellular Responses to Antibody-Toxin Fusion Proteins.

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    Antonella Antignani

    Full Text Available The intersection of small molecular weight drugs and antibody-based therapeutics is rarely studied in large scale. Both types of agents are currently part of the cancer armamentarium. However, very little is known about how to combine them in optimal ways. Immunotoxins are antibody-toxin gene fusion proteins engineered to target cancer cells via antibody binding to surface antigens. For fusion proteins derived from Pseudomonas exotoxin (PE, potency relies on the enzymatic domain of the toxin which catalyzes the ADP-ribosylation of EF2 causing inhibition of protein synthesis leading to cell death. Candidate immunotoxins have demonstrated clear value in clinical trials but generally have not been curative as single agents. Therefore we undertook three screens to discover effective combinations that could act synergistically. From the MIPE-3 library of compounds we identified various enhancers of immunotoxin action and at least one major class of inhibitor. Follow-up experiments confirmed the screening data and suggested that immunotoxins when administered with everolimus or nilotinib exhibit favorable combinatory activity and would be candidates for preclinical development. Mechanistic studies revealed that everolimus-immunotoxin combinations acted synergistically on elements of the protein synthetic machinery, including S61 kinase and 4E-BP1 of the mTORC1 pathway. Conversely, PARP inhibitors antagonized immunotoxins and also blocked the toxicity due to native ADP-ribosylating toxins. Thus, our goal of investigating a chemical library was justified based on the identification of several approved compounds that could be developed preclinically as 'enhancers' and at least one class of mitigator to be avoided.

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

    Science.gov (United States)

    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.

  4. Identifying the Conditions Under Which Antibodies Protect Against Infection by Equine Infectious Anemia Virus.

    Science.gov (United States)

    Schwartz, Elissa J; Smith, Robert J

    2014-05-27

    The ability to predict the conditions under which antibodies protect against viral infection would transform our approach to vaccine development. A more complete understanding is needed of antibody protection against lentivirus infection, as well as the role of mutation in resistance to an antibody vaccine. Recently, an example of antibody-mediated vaccine protection has been shown via passive transfer of neutralizing antibodies before equine infectious anemia virus (EIAV) infection of horses with severe combined immunodeficiency (SCID). Viral dynamic modeling of antibody protection from EIAV infection in SCID horses may lead to insights into the mechanisms of control of infection by antibody vaccination. In this work, such a model is constructed in conjunction with data from EIAV infection of SCID horses to gain insights into multiple strain competition in the presence of antibody control. Conditions are determined under which wild-type infection is eradicated with the antibody vaccine. In addition, a three-strain competition model is considered in which a second mutant strain may coexist with the first mutant strain. The conditions that permit viral escape by the mutant strains are determined, as are the effects of variation in the model parameters. This work extends the current understanding of competition and antibody control in lentiviral infection, which may provide insights into the development of vaccines that stimulate the immune system to control infection effectively.

  5. Identifying the Conditions Under Which Antibodies Protect Against Infection by Equine Infectious Anemia Virus

    Directory of Open Access Journals (Sweden)

    Elissa J. Schwartz

    2014-05-01

    Full Text Available The ability to predict the conditions under which antibodies protect against viral infection would transform our approach to vaccine development. A more complete understanding is needed of antibody protection against lentivirus infection, as well as the role of mutation in resistance to an antibody vaccine. Recently, an example of antibody-mediated vaccine protection has been shown via passive transfer of neutralizing antibodies before equine infectious anemia virus (EIAV infection of horses with severe combined immunodeficiency (SCID. Viral dynamic modeling of antibody protection from EIAV infection in SCID horses may lead to insights into the mechanisms of control of infection by antibody vaccination. In this work, such a model is constructed in conjunction with data from EIAV infection of SCID horses to gain insights into multiple strain competition in the presence of antibody control. Conditions are determined under which wild-type infection is eradicated with the antibody vaccine. In addition, a three-strain competition model is considered in which a second mutant strain may coexist with the first mutant strain. The conditions that permit viral escape by the mutant strains are determined, as are the effects of variation in the model parameters. This work extends the current understanding of competition and antibody control in lentiviral infection, which may provide insights into the development of vaccines that stimulate the immune system to control infection effectively.

  6. Antiphosphatidylserine/prothrombin antibodies as biomarkers to identify severe primary antiphospholipid syndrome.

    Science.gov (United States)

    Hoxha, Ariela; Mattia, Elena; Tonello, Marta; Grava, Chiara; Pengo, Vittorio; Ruffatti, Amelia

    2017-05-01

    Anti-phosphatidylserine/prothrombin (aPS/PT) antibodies have begun to be considered potentional biomarkers for antiphospholipid syndrome (APS). This cohort study investigate the role of aPS/PT antibodies as a risk factor for severe APS by evaluating the association between those antibodies and clinical/laboratory profiles of APS. Plasma/serum samples from 197 APS patients, 100 healthy subjects and 106 patients with autoimmune diseases were collected. IgG/IgM aPS/PT antibodies were assayed using commercial ELISA kit. Prevalences of IgG and IgM aPS/PT (pantiphospholipid antibody patients than in double and single positivity ones (p<0.0001 for all). APS/PT antibodies were associated to severe thrombosis, severe pregnancy complications inducing prematurity, and vascular microangiopathy, all generally associated to high risk APS forms requiring strong therapy.

  7. Use of Moraxella catarrhalis lipooligosaccharide mutants to identify specific oligosaccharide epitopes recognized by human serum antibodies.

    Science.gov (United States)

    Schwingel, Johanna M; Edwards, Katie J; Cox, Andrew D; Masoud, Hussein; Richards, James C; St Michael, Frank; Tekwe, Carmen D; Sethi, Sanjay; Murphy, Timothy F; Campagnari, Anthony A

    2009-10-01

    Moraxella catarrhalis is a causative agent of otitis media in children and lower respiratory tract infections in adults suffering from chronic obstructive pulmonary disease (COPD). This strict human pathogen continues to be a significant cause of disease in this broad spectrum of patients because there is no available vaccine. Although numerous putative vaccine antigens have been described, little is known about the human immune response to M. catarrhalis infection in vivo. Human serum antibodies are directed at a number of surface proteins, and lipooligosaccharides (LOS) and detoxified LOS may be an effective immunogen in mice. In this study, we used a specific LOS-based enzyme-linked immunosorbent assay (ELISA), containing the three major M. catarrhalis serotypes together with a complete series of truncated LOS mutants, to detect the development of new antibodies to specific regions of the oligosaccharide molecule. We compared serum samples from COPD patients who had recently cleared an M. catarrhalis infection to serum samples collected prior to their infection. Variability in the antibody response to LOS was observed, as some patients developed serotype-specific antibodies, others developed antibodies to the LOS of each serotype, others developed broadly cross-reactive antibodies, and some did not develop new antibodies. These newly developed human antibodies are directed at both side chains and core structures in the LOS molecule. This LOS-based ELISA can be used to dissect the human antibody response to both internal and external carbohydrate epitopes, thus providing a better understanding of the humoral immune response to M. catarrhalis LOS epitopes developed during natural infection.

  8. Anti-p-benzoquinone antibody level as a prospective biomarker to identify smokers at risk for COPD

    Directory of Open Access Journals (Sweden)

    Banerjee S

    2017-06-01

    .935 for identifying smokers with COPD from their low antibody level. The antibody cutoff value of 29.4 mg/dL was constructed from the ROC coordinates to estimate the risk for COPD in smokers. While 90.3% of smokers with COPD had a low antibody value (≤29.4 mg/dL, the majority (86.4% of smokers without COPD had a high antibody value (>29.4 mg/dL; 13.6% of current smokers without COPD having an antibody level below this cutoff value (odds ratio [OR] =59.3, 95% CI: 34.15–101.99 were considered to be at risk for COPD.Conclusion and future directions: Our results indicate that serum anti-p-BQ antibody level may be used as a biomarker to identify asymptomatic smokers at risk for COPD for early intervention of the disease. Keywords: COPD, cigarette smoke, biomarker, anti-p-benzoquinone antibody, enzyme-linked immunosorbent assay (ELISA 

  9. Pneumocystis carinii and specific fungi have a common epitope, identified by a monoclonal antibody

    DEFF Research Database (Denmark)

    Lundgren, B; Kovacs, J A; Nelson, N N

    1992-01-01

    Because Pneumocystis carinii may be related to fungi, we evaluated the reactivities of monoclonal antibodies raised against P. carinii with a variety of fungi. Fifty-two fungi and six protozoa were evaluated by immunofluorescence. One of three monoclonal antibodies (MAbs) tested (MAb 7D7) reacted...... with 15 fungi but no protozoa. Saccharomyces cerevisiae showed the strongest reactivity by immunofluorescence. The reactive antigen was characterized for four fungi by the immunoblot technique. In all cases the antigen that was reactive with MAb 7D7 was larger than the P. carinii antigens that reacted...... with 7D7. In further studies with P. carinii, Aspergillus species, and S. cerevisiae, we found that MAb 7D7 reacted with a carbohydrate component in all organisms. The presence of an epitope that is common to P. carinii and a number of fungi further supports the fungal nature of P. carinii....

  10. Spent fuel transport and storage system for NOK: The TN52L, TN97L, TN24 BHL and TN24 GB casks

    International Nuclear Information System (INIS)

    Wattez, L.; Verdier, A.; Monsigny, P.-A.

    2007-01-01

    NOK nuclear power plants in Switzerland, LEIBSTADT (KKL) BWR nuclear power plant and BEZNAU (KKB) PWR nuclear power plant have opted to ship spent fuel to a central facility called ZWILAG for interim storage. In the mid-nineties, COGEMA LOGISTICS was contracted by KKL for the supply of the TN52L and TN97L transport and storage casks for BWR fuel types. In 2003, KKL also ordered from COGEMA LOGISTICS the supply of six TN24 BHL transport and storage casks. This paper shows how all the three cask designs have responded to the KKL needs to ship and store BWR spent fuel. In addition, it highlights the already significant operational feedback of the TN52L and TN97L casks by the KKL and ZWILAG operators. In 2004, NOK also ordered three TN24 GB transport and storage casks for PWR fuel types. These casks are presently being manufactured. (author)

  11. Tennessee Comptroller of the Treasury (TN COMP)

    Data.gov (United States)

    Social Security Administration — The Tennessee Comptroller of the Treasury (TN COMP) uses SSA's assistance in administering the Tennessee Property Tax Rebate Program. SSA provides assistance through...

  12. Epitope Sequences in Dengue Virus NS1 Protein Identified by Monoclonal Antibodies

    Directory of Open Access Journals (Sweden)

    Leticia Barboza Rocha

    2017-10-01

    Full Text Available Dengue nonstructural protein 1 (NS1 is a multi-functional glycoprotein with essential functions both in viral replication and modulation of host innate immune responses. NS1 has been established as a good surrogate marker for infection. In the present study, we generated four anti-NS1 monoclonal antibodies against recombinant NS1 protein from dengue virus serotype 2 (DENV2, which were used to map three NS1 epitopes. The sequence 193AVHADMGYWIESALNDT209 was recognized by monoclonal antibodies 2H5 and 4H1BC, which also cross-reacted with Zika virus (ZIKV protein. On the other hand, the sequence 25VHTWTEQYKFQPES38 was recognized by mAb 4F6 that did not cross react with ZIKV. Lastly, a previously unidentified DENV2 NS1-specific epitope, represented by the sequence 127ELHNQTFLIDGPETAEC143, is described in the present study after reaction with mAb 4H2, which also did not cross react with ZIKV. The selection and characterization of the epitope, specificity of anti-NS1 mAbs, may contribute to the development of diagnostic tools able to differentiate DENV and ZIKV infections.

  13. Multiplex flow cytometry barcoding and antibody arrays identify surface antigen profiles of primary and metastatic colon cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Kumar Sukhdeo

    Full Text Available Colon cancer is a deadly disease affecting millions of people worldwide. Current treatment challenges include management of disease burden as well as improvements in detection and targeting of tumor cells. To identify disease state-specific surface antigen signatures, we combined fluorescent cell barcoding with high-throughput flow cytometric profiling of primary and metastatic colon cancer lines (SW480, SW620, and HCT116. Our multiplexed technique offers improvements over conventional methods by permitting the simultaneous and rapid screening of cancer cells with reduced effort and cost. The method uses a protein-level analysis with commercially available antibodies on live cells with intact epitopes to detect potential tumor-specific targets that can be further investigated for their clinical utility. Multiplexed antibody arrays can easily be applied to other tumor types or pathologies for discovery-based approaches to target identification.

  14. High Concentrations of Measles Neutralizing Antibodies and High-Avidity Measles IgG Accurately Identify Measles Reinfection Cases

    Science.gov (United States)

    Rota, Jennifer S.; Hickman, Carole J.; Mercader, Sara; Redd, Susan; McNall, Rebecca J.; Williams, Nobia; McGrew, Marcia; Walls, M. Laura; Rota, Paul A.; Bellini, William J.

    2016-01-01

    In the United States, approximately 9% of the measles cases reported from 2012 to 2014 occurred in vaccinated individuals. Laboratory confirmation of measles in vaccinated individuals is challenging since IgM assays can give inconclusive results. Although a positive reverse transcription (RT)-PCR assay result from an appropriately timed specimen can provide confirmation, negative results may not rule out a highly suspicious case. Detection of high-avidity measles IgG in serum samples provides laboratory evidence of a past immunologic response to measles from natural infection or immunization. High concentrations of measles neutralizing antibody have been observed by plaque reduction neutralization (PRN) assays among confirmed measles cases with high-avidity IgG, referred to here as reinfection cases (RICs). In this study, we evaluated the utility of measuring levels of measles neutralizing antibody to distinguish RICs from noncases by receiver operating characteristic curve analysis. Single and paired serum samples with high-avidity measles IgG from suspected measles cases submitted to the CDC for routine surveillance were used for the analysis. The RICs were confirmed by a 4-fold rise in PRN titer or by RT-quantitative PCR (RT-qPCR) assay, while the noncases were negative by both assays. Discrimination accuracy was high with serum samples collected ≥3 days after rash onset (area under the curve, 0.953; 95% confidence interval [CI], 0.854 to 0.993). Measles neutralizing antibody concentrations of ≥40,000 mIU/ml identified RICs with 90% sensitivity (95% CI, 74 to 98%) and 100% specificity (95% CI, 82 to 100%). Therefore, when serological or RT-qPCR results are unavailable or inconclusive, suspected measles cases with high-avidity measles IgG can be confirmed as RICs by measles neutralizing antibody concentrations of ≥40,000 mIU/ml. PMID:27335386

  15. Flow cytometry and monoclonal antibodies identify normal liver cell populations antigenically related to oval cells.

    Science.gov (United States)

    Agelli, M; Halay, E D

    1995-01-01

    Oval cells, a non-parenchymal cell population induced to rapidly proliferate in animals treated with carcinogens, are thought to be related to the hypothesized liver stem cells. In normal liver there are poorly defined cells antigenically related to oval cells. These oval cell antigen positive (OCAP) cells present in normal animals are thought to include hepatocyte and bile duct cell precursors. To isolate them, we modified the existing protocols designed for oval cells and used it on normal neonatal rat livers. Using flow cytometry, the percentage of normal liver OCAP-cells varied with the monoclonal antibody (MoAb) to the different oval cell membrane markers used: 12% (MoAb 18.2), 23% (MoAb 270.38), 27% (MoAb 18.11), 31% (MoAb 18.13), and 37% (MoAb 374.3). Macrophages consisted 10% of the cells (MoAb MCA 275); hepatocytes were essentially absent ( < 1%, MoAb 236.4). Our results demonstrate that is possible to obtain significant numbers of normal cells antigenically related to oval cells and that using different MoAbs, different cell populations can be sorted for use in experimental studies testing liver progenitor cell hypothesis.

  16. Prevalence of diphtheria and tetanus antibodies among adults in Singapore: a national serological study to identify most susceptible population groups.

    Science.gov (United States)

    Ang, L W; James, L; Goh, K T

    2016-03-01

    In view of waning antitoxin titres over time after the last vaccine dose against diphtheria and tetanus, we determined the immunity levels in adults to identify most susceptible groups for protection in Singapore. Our study involved residual sera from 3293 adults aged 18-79 who had participated in a national health survey in 2010. IgG antibody levels were determined using commercial enzyme-linked immunosorbent assay. Overall, 92.0% (95% confidence interval [CI]: 91.1-92.9%) had at least basic protection against diphtheria (antibody levels ≥0.01 IU/ml), while 71.4% (95% CI: 69.8-72.9%) had at least short-term protection against tetanus (antibody levels >0.1 IU/ml). The seroprevalence declined significantly with age for both diseases; the drop was most marked in the 50- to 59-year age group for diphtheria and 60- to 69-year age group for tetanus. There was a significant difference in seroprevalence by residency for diphtheria (92.8% among Singapore citizens versus 87.1% among permanent residents; P = 0.001). The seroprevalence for tetanus was significantly higher among males (83.2%) than females (62.4%) (P < 0.0005). It may be of value to consider additional vaccination efforts to protect older adults at higher risk for exposure against diphtheria and tetanus, particularly those travelling to areas where diphtheria is endemic or epidemic. © The Author 2015. Published by Oxford University Press on behalf of Faculty of Public Health. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  17. EnviroAtlas - Memphis, TN - Block Groups

    Data.gov (United States)

    U.S. Environmental Protection Agency — This EnviroAtlas dataset is the base layer for the Memphis, TN EnviroAtlas community. The block groups are from the US Census Bureau and are included/excluded based...

  18. A monoclonal antibody which blocks infection with feline immunodeficiency virus identifies a possible non-CD4 receptor.

    OpenAIRE

    Hosie, M J; Willett, B J; Dunsford, T H; Jarrett, O; Neil, J C

    1993-01-01

    Monoclonal antibody vpg15 detects a 24-kDa cell surface protein on feline cells permissive for infection with feline immunodeficiency virus (FIV). The antibody blocks infection of FIV-susceptible cells, and expression of the vpg15 marker is decreased in FIV-infected cells in vitro. These results suggest that the antibody may recognize an FIV receptor distinct from CD4.

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

    International Nuclear Information System (INIS)

    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

  20. Identifying risk factors for exposure to culturable allergenic moulds in energy efficient homes by using highly specific monoclonal antibodies

    International Nuclear Information System (INIS)

    Sharpe, Richard A.; Cocq, Kate Le; Nikolaou, Vasilis; Osborne, Nicholas J.; Thornton, Christopher R.

    2016-01-01

    The aim of this study was to determine the accuracy of monoclonal antibodies (mAbs) in identifying culturable allergenic fungi present in visible mould growth in energy efficient homes, and to identify risk factors for exposure to these known allergenic fungi. Swabs were taken from fungal contaminated surfaces and culturable yeasts and moulds isolated by using mycological culture. Soluble antigens from cultures were tested by ELISA using mAbs specific to the culturable allergenic fungi Aspergillus and Penicillium spp., Ulocladium, Alternaria, and Epicoccum spp., Cladosporium spp., Fusarium spp., and Trichoderma spp. Diagnostic accuracies of the ELISA tests were determined by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2-encoding regions of recovered fungi following ELISA. There was 100% concordance between the two methods, with ELISAs providing genus-level identity and ITS sequencing providing species-level identities (210 out of 210 tested). Species of Aspergillus/Penicillium, Cladosporium, Ulocladium/Alternaria/Epicoccum, Fusarium and Trichoderma were detected in 82% of the samples. The presence of condensation was associated with an increased risk of surfaces being contaminated by Aspergillus/Penicillium spp. and Cladosporium spp., whereas moisture within the building fabric (water ingress/rising damp) was only associated with increased risk of Aspergillus/Penicillium spp. Property type and energy efficiency levels were found to moderate the risk of indoor surfaces becoming contaminated with Aspergillus/Penicillium and Cladosporium which in turn was modified by the presence of condensation, water ingress and rising damp, consistent with previous literature. - Highlights: • Monoclonal antibodies were used to track culturable allergenic moulds in homes. • Allergenic moulds were recovered from 82% of swabs from contaminated surfaces. • The mAbs were highly specific with 100% agreement to PCR of recovered fungi. • Improvements to energy

  1. Use of epitope libraries to identify exon-specific monoclonal antibodies for characterization of altered dystrophins in muscular dystrophy

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen thi Man; Morris, G.E. (North East Wales Inst., Clwyd (United Kingdom))

    1993-06-01

    The majority of mutations in Xp21-linked muscular dystrophy (MD) can be identified by PCR or Southern blotting, as deletions or duplications of groups of exons in the dystrophin gene, but it is not always possible to predict how much altered dystrophin, if any, will be produced. Use of exon-specific monoclonal antibodies (mAbs) on muscle biopsies from MD patients can, in principle, provide information on both the amount of altered dystrophin produced and, when dystrophin is present, the nature of the genetic deletion or point mutation. For this purpose, mAbs which recognize regions of dystrophin encoded by known exons and whose binding is unaffected by the absence of adjacent exons are required. To map mAbs to specific exons, random [open quotes]libraries[close quotes] of expressed dystrophin fragments were created by cloning DNAseI digestion fragments of a 4.3-kb dystrophin cDNA into a pTEX expression vector. The libraries were then used to locate the epitopes recognized by 48 mAbs to fragments of 25--60 amino acids within the 1,434-amino-acid dystrophin fragment used to produce the antibodies. This is sufficiently detailed to allow further refinement by using synthetic peptides and, in many cases, to identify the exon in the DMD (Duchenne MD) gene which encodes the epitope. To illustrate their use in dystrophin analysis, a Duchenne patient with a frameshift deletion of exons 42 and 43 makes a truncated dystrophin encoded by exons 1--41, and the authors now show that this can be detected in the sarcolemma by mAbs up to and including those specific for exon 41 epitopes but not by mAbs specific for exon 43 or later epitopes. 38 refs., 2 figs., 4 tabs.

  2. Proteomic profiling of antibody-inducing immunogens in tumor tissue identifies PSMA1, LAP3, ANXA3, and maspin as colon cancer markers

    Science.gov (United States)

    Yang, Qian; Roehrl, Michael H.; Wang, Julia Y.

    2018-01-01

    We hypothesized that cancer tissue immunogens – antigens capable of inducing specific antibody production in patients – are promising targets for development of precision diagnostics and humoral immunotherapies. We developed an innovative immuno-proteomic strategy and identified new immunogenic markers of colon cancer. Proteins from cancers and matched normal tissues were separated by 2D gel electrophoresis and blotted with serum antibodies from the same patients. Antibody-reactive proteins were sequenced by mass spectrometry and validated by Western blotting and immunohistochemistry. 170 serum antibody-reactive proteins were identified only in cancerous but not matched normal. Among these, proteasome subunit alpha type 1 (PSA1), leucine aminopeptidase 3 (LAP3), annexin A3 (ANXA3), and maspin (serpin B5) were reproducibly found in tissues from three patients. Differential expression patterns were confirmed in samples from eight patients with various stages of colon adenocarcinoma and liver metastases. These tumor-resident proteins and/or their associated serum antibodies may be promising markers for colon cancer screening and early diagnosis. Furthermore, tumor tissue-specific antibodies could potentially be exploited as immunotherapeutic targets against cancer. More generally, proteomic profiling of antibody-inducing cancer-associated immunogens represents a powerful generic method for uncovering the tumor antigen-ome, i.e., the totality of immunogenic tumor-associated proteins. PMID:29423100

  3. A novel ex vivo immunoproteomic approach characterising Fasciola hepatica tegumental antigens identified using immune antibody from resistant sheep.

    Science.gov (United States)

    Cameron, Timothy C; Cooke, Ira; Faou, Pierre; Toet, Hayley; Piedrafita, David; Young, Neil; Rathinasamy, Vignesh; Beddoe, Travis; Anderson, Glenn; Dempster, Robert; Spithill, Terry W

    2017-08-01

    A more thorough understanding of the immunological interactions between Fasciola spp. and their hosts is required if we are to develop new immunotherapies to control fasciolosis. Deeper knowledge of the antigens that are the target of the acquired immune responses of definitive hosts against both Fasciola hepatica and Fasciola gigantica will potentially identify candidate vaccine antigens. Indonesian Thin Tail sheep express a high level of acquired immunity to infection by F. gigantica within 4weeks of infection and antibodies in Indonesian Thin Tail sera can promote antibody-dependent cell-mediated cytotoxicity against the surface tegument of juvenile F. gigantica in vitro. Given the high protein sequence similarity between F. hepatica and F. gigantica, we hypothesised that antibody from F. gigantica-infected sheep could be used to identify the orthologous proteins in the tegument of F. hepatica. Purified IgG from the sera of F. gigantica-infected Indonesian Thin Tail sheep collected pre-infection and 4weeks p.i. were incubated with live adult F. hepatica ex vivo and the immunosloughate (immunoprecipitate) formed was isolated and analysed via liquid chromatography-electrospray ionisation-tandem mass spectrometry to identify proteins involved in the immune response. A total of 38 proteins were identified at a significantly higher abundance in the immunosloughate using week 4 IgG, including eight predicted membrane proteins, 20 secreted proteins, nine proteins predicted to be associated with either the lysosomes, the cytoplasm or the cytoskeleton and one protein with an unknown cellular localization. Three of the membrane proteins are transporters including a multidrug resistance protein, an amino acid permease and a glucose transporter. Interestingly, a total of 21 of the 38 proteins matched with proteins recently reported to be associated with the proposed small exosome-like extracellular vesicles of adult F. hepatica, suggesting that the Indonesian Thin Tail week

  4. A novel merozoite surface antigen of Plasmodium falciparum (MSP-3 identified by cellular-antibody cooperative mechanism antigenicity and biological activity of antibodies

    Directory of Open Access Journals (Sweden)

    Claude Oeuvray

    1994-01-01

    Full Text Available We report the identification of a 48kDa antigen targeted by antibodies which inhibit Plasmodium falciparum in vitro growth by cooperation with blood monocytes in an ADCI assay correlated to the naturally acquired protection. This protein is located on the surface of the merozoite stage of P. falciparum, and is detectable in all isolates tested. Epidemiological studies demonstrated that peptides derived from the amino acid sequence of MSP-3 contain potent B and T-cell epitopes recognized by a majority of individuals living in endemic areas. Moreover human antibodies either purified on the recombinant protein, or on the synthetic peptide MSP-3b, as well as antibodies raised in mice, were all found to promote parasite killing mediated by monocytes.

  5. Rabbit monoclonal antibodies directed at the T3SS effector protein YopM identify human pathogenic Yersinia isolates.

    Science.gov (United States)

    Rüter, Christian; Silva, Mariana Ruiz; Grabowski, Benjamin; Lubos, Marie-Luise; Scharnert, Julia; Poceva, Marija; von Tils, Dominik; Flieger, Antje; Heesemann, Jürgen; Bliska, James B; Schmidt, M Alexander

    2014-05-01

    The Yersinia outer protein M (YopM) is a type 3 secretion system (T3SS)-dependent effector protein of Yersinia enterocolitica, Yersinia pseudotuberculosis and Yersinia pestis. Although YopM is indispensable for full virulence, its molecular functions still remain largely elusive. Recently, we could identify the recombinant YopM (rYopM) protein derived from the Y. enterocolitica strain 8081 (JB580) as a cell-penetrating protein, which down-regulates the expression of various pro-inflammatory cytokines including TNFα. In this study, we have generated rabbit monoclonal anti-YopM antibodies (RabMabs). RabMabs were characterized by SDS-PAGE and Western blotting using various truncated versions of rYopM to identify epitope-containing domains. RabMabs recognizing either the N- or C-terminus of YopM were characterized further and validated using a collection of 61 pathogenic and non-pathogenic Yersinia strains as well as exemplary strains of major intestinal bacterial pathogens such as Salmonella enterica ssp. enterica, Shigella flexneri and intestinal pathogenic Escherichia coli. RabMab 41.3 directed at the N-terminus of YopM of Y. enterocolitica strain 8081 recognized all YopM-expressing pathogenic Yersinia strains analyzed in this study but failed to recognize non-pathogenic isolates. Thus, RabMab 41.3 might be applicable for the detection of pathogenic Yersinia strains. Copyright © 2014 Elsevier GmbH. All rights reserved.

  6. A clinical isolate of transposon Tn5 expressing streptomycin resistance in Escherichia coli.

    OpenAIRE

    Genilloud, O; Blázquez, J; Mazodier, P; Moreno, F

    1988-01-01

    The central region of transposon Tn5 carries three antibiotic resistance markers: neo, ble, and str. The str gene codes for a phosphotransferase that inactivates streptomycin. This activity is phenotypically expressed in several gram-negative bacteria but not in Escherichia coli. We identified a Tn5 variant in E. coli clinical isolates that express streptomycin resistance. This transposon carries a 6-base-pair deletion within the str gene, near the 3' end. The same kind of mutation had been p...

  7. Genetic and molecular studies of a composite chromosomal element (Tn3705) containing a Tn916-modified structure (Tn3704) in Streptococcus anginosus F22.

    Science.gov (United States)

    Clermont, D; Horaud, T

    1994-01-01

    The plasmid-free Streptococcus anginosus F22 contained a conjugative element, Tn3705, encoding resistance to erythromycin (Emr) and tetracycline-minocycline (Tcr-Mnr). We mapped a chromosomal region (> 52 kb) of F22, corresponding to the internal part of Tn3705. Molecular analysis of Tn3705 revealed it to be a composite structure: it included in its central part a transposon designated Tn3704 (20.3 kb +/- 0.5 kb), which had a modified structure in comparison with that of Tn916 and on which the Emr Tcr-Mn4 markers were localized. Tn3705 inserted from F22 into the chromosome of various streptococcal transconjugants as well as that of Enterococcus faecalis transconjugants without changing its structure. In contrast, from the chromosome of an E. faecalis::Tn3705 transconjugant only Tn3704 inserted, at various sites, into another E. faecalis chromosome. Sugar fermentations occurred after the insertion of Tn3704 into the chromosome of an asaccharolytic E. faecalis strain. Transposition of only Tn3704 from the chromosome of E. faecalis::Tn3705 onto pIP964, an E. faecalis hemolysin plasmid, yielded two different pIP964 derivatives. The size of the entire element Tn3705 was estimated to be about 70.0 kb by pulsed-field electrophoresis.

  8. Antibody responses to NY-ESO-1 in primary breast cancer identify a subtype target for immunotherapy.

    Science.gov (United States)

    Hamaï, Ahmed; Duperrier-Amouriaux, Karine; Pignon, Pascale; Raimbaud, Isabelle; Memeo, Lorenzo; Colarossi, Cristina; Canzonieri, Vincenzo; Perin, Tiziana; Classe, Jean-Marc; Campone, Mario; Jézéquel, Pascal; Campion, Loïc; Ayyoub, Maha; Valmori, Danila

    2011-01-01

    The highly immunogenic human tumor antigen NY-ESO-1 (ESO) is a target of choice for anti-cancer immune therapy. In this study, we assessed spontaneous antibody (Ab) responses to ESO in a large cohort of patients with primary breast cancer (BC) and addressed the correlation between the presence of anti-ESO Ab, the expression of ESO in the tumors and their characteristics. We found detectable Ab responses to ESO in 1% of the patients. Tumors from patients with circulating Ab to ESO exhibited common characteristics, being mainly hormone receptor (HR)⁻ invasive ductal carcinomas of high grade, including both HER2⁻ and HER2⁺ tumors. In line with these results, we detected ESO expression in 20% of primary HR⁻ BC, including both ESO Ab⁺ and Ab⁻ patients, but not in HR⁺ BC. Interestingly, whereas expression levels in ESO⁺ BC were not significantly different between ESO Ab⁺ and Ab⁻ patients, the former had, in average, significantly higher numbers of tumor-infiltrated lymph nodes, indicating that lymph node invasion may be required for the development of spontaneous anti-tumor immune responses. Thus, the presence of ESO Ab identifies a tumor subtype of HR⁻ (HER2⁻ or HER2⁺) primary BC with frequent ESO expression and, together with the assessment of antigen expression in the tumor, may be instrumental for the selection of patients for whom ESO-based immunotherapy may complement standard therapy.

  9. The fully synthetic MAG-Tn3 therapeutic vaccine containing the tetanus toxoid-derived TT830-844 universal epitope provides anti-tumor immunity.

    Science.gov (United States)

    Laubreton, Daphné; Bay, Sylvie; Sedlik, Christine; Artaud, Cécile; Ganneau, Christelle; Dériaud, Edith; Viel, Sophie; Puaux, Anne-Laure; Amigorena, Sebastian; Gérard, Catherine; Lo-Man, Richard; Leclerc, Claude

    2016-03-01

    Malignant transformations are often associated with aberrant glycosylation processes that lead to the expression of new carbohydrate antigens at the surface of tumor cells. Of these carbohydrate antigens, the Tn antigen is particularly highly expressed in many carcinomas, especially in breast carcinoma. We designed MAG-Tn3, a fully synthetic vaccine based on three consecutive Tn moieties that are O-linked to a CD4+ T cell epitope, to induce anti-Tn antibody responses that could be helpful for therapeutic vaccination against cancer. To ensure broad coverage within the human population, the tetanus toxoid-derived peptide TT830-844 was selected as a T-helper epitope because it can bind to various HLA-DRB molecules. We showed that the MAG-Tn3 vaccine, which was formulated with the GSK proprietary immunostimulant AS15 and designed for human cancer therapy, is able to induce an anti-Tn antibody response in mice of various H-2 haplotypes, and this response correlates with the ability to induce a specific T cell response against the TT830-844 peptide. The universality of the TT830-844 peptide was extended to new H-2 and HLA-DRB molecules that were capable of binding this T cell epitope. Finally, the MAG-Tn3 vaccine was able to induce anti-Tn antibody responses in cynomolgus monkeys, which targeted Tn-expressing tumor cells and mediated tumor cell death both in vitro and in vivo. Thus, MAG-Tn3 is a highly promising anticancer vaccine that is currently under evaluation in a phase I clinical trial.

  10. Identifying long-term memory B-cells in vaccinated children despite waning antibody levels specific for Bordetella pertussis proteins

    NARCIS (Netherlands)

    Hendrikx, Lotte H.; Ozturk, Kemal; de Rond, Lia G. H.; Veenhoven, Reinier H.; Sanders, Elisabeth A. M.; Berbers, Guy A. M.; Buisman, Anne-Marie

    2011-01-01

    Whooping cough is a respiratory disease caused by Bordetella pertussis. Since the 1950s in developed countries pertussis vaccinations are included in the national immunization program. However, antibody levels rapidly wane after both whole cell and acellular pertussis vaccination. Therefore

  11. [Construction of cTnC-linker-TnI (P) Genes, Expression of Fusion Protein and Preparation of Lyophilized Protein].

    Science.gov (United States)

    Song, Xiaoli; Liu, Xiaoyun; Cai, Lei; Wu, Jianwei; Wang, Jihua

    2015-12-01

    In order to construct and express human cardiac troponin C-linker-troponin I(P) [ cTnC-linker-TnI(P)] fusion protein, detect its activity and prepare lyophilized protein, we searched the CDs of human cTnC and cTnI from GenBank, synthesized cTnC and cTnI(30-110aa) into cloning vector by a short DNA sequence coding for 15 neutral amino acid residues. pCold I-cTnC-linker-TnI(P) was constructed and transformed into E. coli BL21(DE3). Then, cTnC-linker-TnI(P) fusion protein was induced by isopropyl-β-D-thiogalactopyranoside (IPTG). Soluable expression of cTnC-linker-TnI(P) in prokaryotic system was successfully obtained. The fusion protein was purified by Ni²⁺ Sepharose 6 Fast Flow affinity chromatography with over 95% purity and prepared into lyophilized protein. The activity of purified cTnC-linker-TnI(P) and its lyophilized protein were detected by Wondfo Finecare™ cTnI Test. Lyophilized protein of cTnC-linker-TnI(P) was stable for 10 or more days at 37 °C and 4 or more months at 25 °C and 4 °C. The expression system established in this research is feasible and efficient. Lyophilized protein is stable enough to be provided as biological raw materials for further research.

  12. Field validation of a commercial blocking ELISA to differentiate antibody to transmissible gastroenteritis virus (TGEV) and porcine respiratory coronavirus and to identify TGEV-infected swine herds.

    Science.gov (United States)

    Carman, Susy; Josephson, Gaylan; McEwen, Beverly; Maxie, Grant; Antochi, Mioara; Eernisse, Ken; Nayar, Gopi; Halbur, Pat; Erickson, Gene; Nilsson, Ernst

    2002-03-01

    A commercially available blocking ELISA was analyzed for its ability to identify antibodies to porcine coronaviruses (transmissible gastroenteritis virus [TGEV] or porcine respiratory coronavirus [PRCV]), to differentiate antibodies to TGEV and PRCV, and to identify TGEV-infected herds. Nine sera from uninfected pigs, 34 sera from 16 pigs experimentally infected with TGEV, and sera from 10 pigs experimentally infected with PRCV were evaluated using both the TGEV/PRCV blocking ELISA and a virus neutralization (VN) assay. The ELISA was not consistently effective in identifying pigs experimentally infected with TGEV until 21 days postinfection. Sera from 100 commercial swine herds (1,783 sera; median 15 per herd) were similarly evaluated using both tests. Thirty of these commercial herds had a clinical history of TGEV infection and a positive TGEV fluorescent antibody test recorded at necropsy within the last 35 months, while 70 herds had no history of clinical TGEV infection. The blocking ELISA and the VN showed good agreement (kappa 0.84) for the detection of porcine coronavirus antibody (TGEV or PRCV). The sensitivity (0.933) of the ELISA to identify TGEV-infected herds was good when considered on a herd basis. The ELISA was also highly specific (0.943) for the detection of TGEV-infected herds when the test results were evaluated on a herd basis. When sera from specific age groups were compared, the ELISA identified a greater proportion (0.83) of pigs in herds with TGEV antibody when suckling piglets were used. In repeatability experiments, the ELISA gave consistent results when the same sera were evaluated on different days (kappa 0.889) and when sera were evaluated before and after heating (kappa 0.888). The blocking ELISA was determined to be useful for herd monitoring programs and could be used alone without parallel use of the VN assay for the assessment of large swine populations for the detection of TGEV-infected herds.

  13. International Spread and Persistence of TEM-24 Is Caused by the Confluence of Highly Penetrating Enterobacteriaceae Clones and an IncA/C2 Plasmid Containing Tn1696::Tn1 and IS5075-Tn21▿

    Science.gov (United States)

    Novais, Ângela; Baquero, Fernando; Machado, Elisabete; Cantón, Rafael; Peixe, Luísa; Coque, Teresa M.

    2010-01-01

    TEM-24 remains one of the most widespread TEM-type extended-spectrum β-lactamases (ESBLs) among Enterobacteriaceae. To analyze the reasons influencing its spread and persistence, a multilevel population genetics study was carried out on 28 representative TEM-24 producers from Belgium, France, Portugal, and Spain (13 Enterobacter aerogenes isolates, 6 Escherichia coli isolates, 6 Klebsiella pneumoniae isolates, 2 Proteus mirabilis isolates, and 1 Klebsiella oxytoca isolate, from 1998 to 2004). Clonal relatedness (XbaI pulsed-field gel electrophoresis [PFGE] and E. coli phylogroups) and antibiotic susceptibility were determined by standard procedures. Plasmid analysis included determination of the incompatibility group (by PCR, hybridization, and/or sequencing) and comparison of restriction fragment length polymorphism (RFLP) patterns. Characterization of genetic elements conferring antibiotic resistance included integrons (classes 1, 2, and 3) and transposons (Tn3, Tn21, and Tn402). Similar PFGE patterns were identified among E. aerogenes, K. pneumoniae, and P. mirabilis isolates, while E. coli strains were diverse (phylogenetic groups A, B2, and D). Highly related 180-kb IncA/C2 plasmids conferring resistance to kanamycin, tobramycin, chloramphenicol, trimethoprim, and sulfonamides were identified. Each plasmid contained defective In0-Tn402 (dfrA1-aadA1, aacA4, or aacA4-aacC1-orfE-aadA2-cmlA1) and In4-Tn402 (aacA4 or dfrA1-aadA1) variants. These integrons were located within Tn21, Tn1696, or hybrids of these transposons, with IS5075 interrupting their IRtnp and IRmer. In all cases, blaTEM-24 was part of an IS5075-ΔTn1 transposon within tnp1696, mimicking other genetic elements containing blaTEM-2 and blaTEM-3 variants. The international dissemination of TEM-24 is fuelled by an IncA/C2 plasmid acquired by different enterobacterial clones which seem to evolve by gaining diverse genetic elements. This work highlights the risks of a confluence between highly

  14. Simple mucins (T, sialosyl-T, Tn and sialosyl-Tn) are not diagnostic for malignant breast lesions

    DEFF Research Database (Denmark)

    Reed, W; Bryne, M; Clausen, H

    1994-01-01

    carcinomas (79 localized and 43 metastatic lesions). T antigen, not observed in normal breast tissue, was present in 31% of the benign lesions and in some cases of morphologically normal epithelium adjacent to tumor cells, compatible with altered glycosylation being an early event. Sialosyl-T (s-T) antigen...... was present in all cases of normal epithelium and in 81% of the benign lesions. Both Tn and sialosyl-Tn (s-Tn) antigen were present in normal breast lesions. Both Tn and sialosyl-Tn (s-Tn) antigen were present in normal breast tissue (30%) and benign lesions (31% and 19%). In the malignant lesions, 20% were...... positive for T antigen, 82% for s-T antigen, 66% for Tn antigen and 22% for s-Tn antigen. The staining pattern was nearly identical for carcinomas with and without lymph node metastases. In conclusion, immunostaining for simple mucins does not permit a clear distinction between benign and malignant breast...

  15. Tn7 and Tn501 Insertions into Pseudomonas aeruginosa plasmid R91-5: mapping of two transfer regions.

    OpenAIRE

    Moore, R J; Krishnapillai, V

    1982-01-01

    We constructed a restriction endonuclease map of the Pseudomonas aeruginosa narrow-host-range plasmid R91-5. Insertions of transposons Tn7 and Tn501 into the plasmid DNA were characterized physically and genetically. The distribution of sites of insertion showed some regional specificity for the insertion of these transposons, especially TN501. The insertion of Tn7 was unusual in that all 42 of 43 insertions were in the same orientation. By relating phenotypic changes to the site of insertion...

  16. Mechanism for transfer of transposon Tn2010 carrying macrolide resistance genes in Streptococcus pneumoniae and its effects on genome evolution.

    Science.gov (United States)

    Zhou, Wenqing; Yao, Kaihu; Zhang, Gang; Yang, Yonghong; Li, Yun; Lv, Yuan; Feng, Jie

    2014-06-01

    The objective of this study was to identify the mechanism responsible for the horizontal transfer of transposon Tn2010 in Streptococcus pneumoniae, and the genomic alterations introduced by the transfer process. Tn2010 was identified using PCR in 15 clinical isolates of S. pneumoniae with erythromycin resistance. S. pneumoniae and Enterococcus faecalis isolates were used as recipient cells in mating and transformation experiments to test the conjugative transferability and transformability of Tn2010. Whole-genome sequencing was used to assess the effects of the Tn2010 transfer on recipient genomes. The biological cost of the horizontal acquisition of Tn2010 and additional genomic changes was investigated by growth competition experiments. Tn2010 was transformed at a frequency of 3 × 10(-7) transformants per cfu, whereas no transconjugants were detected using S. pneumoniae or E. faecalis as recipient cells. Genome analysis showed that many other recombinations were scattered throughout the genome of the transformants in addition to transposon Tn2010. The transformants demonstrated a negligible fitness cost compared with the wild-type strain. Tn2010 tended to be transferred by transformation rather than conjugation in S. pneumoniae, and the spread of Tn2010 could have a profound effect on the evolution of the genome. The acquisition of Tn2010 with negligible fitness cost may facilitate spread of the transposon. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  17. Control charts for identifying systematic errors using control sera to detect antibody to Salmonella in an indirect ELISA

    DEFF Research Database (Denmark)

    Bak, H.; Barfod, Kristen

    2008-01-01

    This study evaluated the preparation of Shewhart's control charts using the concept of rational subgroups for monitoring the Salmonella antibody ELISA used for surveillance of Danish pig herds. Control charts were prepared for a buffer control sample, a negative serum sample and a positive serum ...

  18. Neuronal antibody biomarkers for Sydenham's chorea identify a new group of children with chronic recurrent episodic acute exacerbations of tic and obsessive compulsive symptoms following a streptococcal infection.

    Directory of Open Access Journals (Sweden)

    Harvey S Singer

    Full Text Available Several autoantibodies (anti-dopamine 1 (D1R and 2 (D2R receptors, anti-tubulin, anti-lysoganglioside-GM1 and antibody-mediated activation of calcium calmodulin dependent protein kinase II (CaMKII signaling activity are elevated in children with Sydenham's chorea (SC. Recognizing proposed clinical and autoimmune similarities between SC and PANDAS (pediatric autoimmune neuropsychiatric disorder associated with a streptococcal infection, we sought to identify serial biomarker changes in a slightly different population. Antineuronal antibodies were measured in eight children (mean 11.3 years with chronic, dramatic, recurrent tics and obsessive-compulsive disorder (OCD associated with a group A β-hemolytic streptococcal (GABHS respiratory tract infection, but differing because they lacked choreiform movements. Longitudinal serum samples in most subjects included two pre-exacerbation samples, Exac, one midst Exac (abrupt recurrence of tic/OCD; temporally association with a GABHS infection in six of eight subjects, and two post-Exac. Controls included four groups of unaffected children (n = 70; mean 10.8 years obtained at four different institutions and published controls. Clinical exacerbations were not associated with a significant rise in antineuronal antibody titers. CaMKII activation was increased at the GABHS exacerbation point in 5/6 subjects, exceeded combined and published control's 95th percentile at least once in 7/8 subjects, and median values were elevated at each time point. Anti-tubulin and anti-D2R titers did not differ from published or combined control group's 95th percentile or median values. Differences in anti-lysoganglioside-GM1 and anti-D1R titers were dependent on the selected control. Variances in antibody titers and CaMKII activation were identified among the institutional control groups. Based on comparisons to published studies, results identify two groups of PANDAS: 1 a cohort, represented by this study, which lacks

  19. Ab-origin: an enhanced tool to identify the sourcing gene segments in germline for rearranged antibodies

    Directory of Open Access Journals (Sweden)

    Sun Jing

    2008-12-01

    Full Text Available Abstract Background In the adaptive immune system, variable regions of immunoglobulin (IG are encoded by random recombination of variable (V, diversity (D, and joining (J gene segments in the germline. Partitioning the functional antibody sequences to their sourcing germline gene segments is vital not only for understanding antibody maturation but also for promoting the potential engineering of the therapeutic antibodies. To date, several tools have been developed to perform such "trace-back" calculations. Yet, the predicting ability and processing volume of those tools vary significantly for different sets of data. Moreover, none of them give a confidence for immunoglobulin heavy diversity (IGHD identification. Developing fast, efficient and enhanced tools is always needed with the booming of immunological data. Results Here, a program named Ab-origin is presented. It is designed by batch query against germline databases based on empirical knowledge, optimized scoring scheme and appropriate parameters. Special efforts have been paid to improve the identification accuracy of the short and volatile region, IGHD. In particular, a threshold score for certain sensitivity and specificity is provided to give the confidence level of the IGHD identification. Conclusion When evaluated using different sets of both simulated data and experimental data, Ab-origin outperformed all the other five popular tools in terms of prediction accuracy. The features of batch query and confidence indication of IGHD identification would provide extra help to users. The program is freely available at http://mpsq.biosino.org/ab-origin/supplementary.html.

  20. Identifying long-term memory B-cells in vaccinated children despite waning antibody levels specific for Bordetella pertussis proteins.

    Science.gov (United States)

    Hendrikx, Lotte H; Oztürk, Kemal; de Rond, Lia G H; Veenhoven, Reinier H; Sanders, Elisabeth A M; Berbers, Guy A M; Buisman, Anne-Marie

    2011-02-04

    Whooping cough is a respiratory disease caused by Bordetella pertussis. Since the 1950s in developed countries pertussis vaccinations are included in the national immunization program. However, antibody levels rapidly wane after both whole cell and acellular pertussis vaccination. Therefore protection against pertussis may depend largely on long-term B- and T-cell immunities. We investigated long-term pertussis-specific memory B-cell responses in children who were primed at infant age with the Dutch wP-vaccine (ISRCTN65428640). Purified B-cells were characterized by FACS-analysis and after polyclonal stimulation memory B-cells were detected by ELISPOT-assays specific for pertussis toxin, filamentous haemagglutinin, pertactin and tetanus. In addition, plasma IgG levels directed to the same antigens were measured by a fluorescent bead-based multiplex immunoassay. Two and 3 years after wP priming as well as 2 and 5 years after the aP booster at the age of 4, low plasma IgG levels to the pertussis proteins were found. At the same time, however pertussis protein-specific memory B-cells could be detected and their number increased with age. The number of tetanus-specific memory B-cells was similar in all age groups, whereas IgG-tetanus levels were high 2 years after tetanus booster compared to pre- and 5 years post-booster levels. This study shows the presence of long-term pertussis protein-specific memory B-cells in children despite waning antibody levels after vaccination, which suggests that memory B-cells in addition to antibodies may contribute to protection against pertussis. Copyright © 2010 Elsevier Ltd. All rights reserved.

  1. An HLA-B27 Homodimer Specific Antibody Recognizes a Discontinuous Mixed-Disulfide Epitope as Identified by Affinity-Mass Spectrometry

    Science.gov (United States)

    Iuraşcu, Marius-Ionuţ; Marroquin Belaunzanar, Osiris; Cozma, Claudia; Petrausch, Ulf; Renner, Christoph; Przybylski, Michael

    2016-06-01

    HLA-B27 homodimer formation is believed to be a hallmark of HLA-B27 associated spondyloarthritides. Recently, we have generated a homodimer-specific monoclonal antibody (HD6) and have demonstrated that HLA-B27 homodimer complexes are present on monocytes of healthy HLA-B27 gene carriers at low levels, with significantly increased levels at active disease. The capability of the HD6 antibody to discriminate between correctly formed HLA-B27 heterotrimers and pathology-associated homodimers is striking and cannot be explained by the primary structure of HLA-B27. We hypothesized that HD6 accesses a unique epitope and used affinity-mass spectrometry for its identification. The HD6 antibody was immobilized on an activated sepharose affinity column, and HLA-B27 homodimer characterized for affinity. The epitope was identified by proteolytic epitope excision and MALDI mass spectrometry, and shown to comprise a discontinuous Cys-203- 257-Cys mixed-disulfide peptide structure that is not accessible in HLA-B27 heterotrimers due to protection by noncovalently linked β2-microglobulin. The epitope peptides were synthesized by solid phase peptide synthesis, and the two monomeric peptide components, HLA-B27(203-219) and HLA-B27(257-273), as well as the homo- and hetero-dimeric disulfide linked combinations prepared. The affinity binding constants KD towards the antibodies were determined using a surface acoustic wave (SAW) biosensor, and showed the highest affinity with a KD of approximately 40 nM to the HD6 antibody for the (203-219)-SS-(257-273) mixed disulfide epitope.

  2. The importance of Foxp3 antibody and fixation/permeabilization buffer combinations in identifying CD4+CD25+Foxp3+ regulatory T cells.

    Science.gov (United States)

    Law, Jacqueline P; Hirschkorn, Dale F; Owen, Rachel E; Biswas, Hope H; Norris, Philip J; Lanteri, Marion C

    2009-12-01

    Foxp3 is a key marker for CD4(+) regulatory T cells (T(regs)) and was used in developing a multiparameter flow cytometric panel to identify T(regs). Achieving reproducible staining and analysis first required optimization of Foxp3 staining. We present a comparative study of PCH101, 236A/E7, 3G3, 206D, 150D, and 259D/C7 clones of anti-human-Foxp3 antibodies used in combination with five different fixation/permeabilization buffers. Staining for CD25, CD152, and CD127 was also compared between fixation/permeabilization treatments. Promising antibody/buffer combinations were tested in a panel of peripheral blood mononuclear cells from 10 individuals, and then on fresh versus frozen cells from four individuals. Finally, different fluorochromes coupled to two representative antibodies were compared to optimize separation of Foxp3(+) from Foxp3(-) events. Foxp3 gates were set using two gating strategies based on CD127(+)CD25(-) "non-T(regs)" or based on isotype controls. For Foxp3 staining, the best conditions for fixation/permeabilization were obtained using the eBioscience Foxp3, Imgenex, BioLegend, and BD Foxp3 buffers. Comparing results from 10 subjects, 259D/C7, PCH101, 236A/E7, and 206D antibodies yielded statistically higher levels of Foxp3 cells than those by 150D and 3G3 antibodies (mean = 6.9, 5.1, 4.7, and 3.7% compared with 1.7, and 0.3% of CD25(+)Foxp3(+) events within CD4(+) cells, respectively). Importantly, the "nonspecificity" of some antibodies observed with a Foxp3 gate based on isotype controls could be eliminated by setting the Foxp3 gate on "non-T(regs)". Better separation of Foxp3(+) and Foxp3(-) populations was observed using the PCH101 clone coupled to Alexa647 compared with FITC or the 259D/C7 clone coupled to PE compared with Alexa488 fluorochrome. Foxp3 staining can be highly variable and depends on the choice of antibody/buffer pair and the fluorochrome used. Selecting the correct population for setting the Foxp3 gate is critical to avoid

  3. TRANSIT--A Software Tool for Himar1 TnSeq Analysis.

    Directory of Open Access Journals (Sweden)

    Michael A DeJesus

    2015-10-01

    Full Text Available TnSeq has become a popular technique for determining the essentiality of genomic regions in bacterial organisms. Several methods have been developed to analyze the wealth of data that has been obtained through TnSeq experiments. We developed a tool for analyzing Himar1 TnSeq data called TRANSIT. TRANSIT provides a graphical interface to three different statistical methods for analyzing TnSeq data. These methods cover a variety of approaches capable of identifying essential genes in individual datasets as well as comparative analysis between conditions. We demonstrate the utility of this software by analyzing TnSeq datasets of M. tuberculosis grown on glycerol and cholesterol. We show that TRANSIT can be used to discover genes which have been previously implicated for growth on these carbon sources. TRANSIT is written in Python, and thus can be run on Windows, OSX and Linux platforms. The source code is distributed under the GNU GPL v3 license and can be obtained from the following GitHub repository: https://github.com/mad-lab/transit.

  4. An Enterobacter plasmid as a new genetic background for the transposon Tn1331.

    Science.gov (United States)

    Alavi, Mohammad R; Antonic, Vlado; Ravizee, Adrien; Weina, Peter J; Izadjoo, Mina; Stojadinovic, Alexander

    2011-01-01

    Genus Enterobacter includes important opportunistic nosocomial pathogens that could infect complex wounds. The presence of antibiotic resistance genes in these microorganisms represents a challenging clinical problem in the treatment of these wounds. In the authors' screening of antibiotic-resistant bacteria from complex wounds, an Enterobacter species was isolated that harbors antibiotic-resistant plasmids conferring resistance to Escherichia coli. The aim of this study was to identify the resistance genes carried by one of these plasmids. The plasmids from the Enterobacter isolate were propagated in E. coli and one of the plasmids, designated as pR23, was sequenced by the Sanger method using fluorescent dyeterminator chemistry on a genetic analyzer. The assembled sequence was annotated by search of the GenBank database. Plasmid pR23 is composed of the transposon Tn1331 and a backbone plasmid that is identical to the plasmid pPIGDM1 from Enterobacter agglomerans. The multidrug-resistance transposon Tn1331, which confers resistance to aminoglycoside and beta lactam antibiotics, has been previously isolated only from Klebsiella. The Enterobacter plasmid pPIGDM1, which carries a ColE1-like origin of replication and has no apparent selective marker, appears to provide a backbone for propagation of Tn1331 in Enterobacter. The recognition sequence of Tn1331 transposase for insertion into pPIGDM1 is the pentanucleotide TATTA, which occurs only once throughout the length of this plasmid. Transposition of Tn1331 into the Enterobacter plasmid pPIGDM1 enables this transposon to propagate in this Enterobacter. Since Tn1331 was previously isolated only from Klebsiella, this report suggests horizontal transfer of this transposon between the two bacterial genera.

  5. Interaction of a novel Tn (GalNAc alpha 1-->Ser/Thr) glycoprotein with Gal, GalNAc and GlcNAc specific lectins.

    Science.gov (United States)

    Wu, A M; Wu, J H; Shen, F

    1994-01-14

    A naturally occurring Tn glycoprotein (Native ASG-Tn) with GalNAc alpha 1-->Ser/Thr as the only carbohydrate side chains, has been prepared from armadillo submandibular glands. In a quantitative precipitin assay, this glycoprotein completely precipitated Maclura pomifera (MPA), Vicia villosa B4 (VVL-B4) and Artocarpus integrifolia (Jacalin, AIL). It also reacted well with Helix pomatia (HPL) and Wistaria floribunda (WFL) and precipitated over 75% of the lectin nitrogen added, but poorly with Ricinus communis agglutinin (RCA1), ricin, peanut (Arachis hypogaea, PNA), Abrus precatorius agglutinin (APA) and Triticum vulgaris (WGA). This finding suggests that this novel Tn-glycoprotein may serve as a useful reagent for differentiating Tn and T specific monoclonal antibodies and lectins.

  6. BWR spent fuel transport and storage system for KKL: TN trademark 52L, TN trademark 97L, TN trademark 24 BHL

    International Nuclear Information System (INIS)

    Sicard, D.; Verdier, A.; Monsigny, P.A.

    2004-01-01

    The LEIBSTADT (KKL) nuclear power plant in Switzerland has opted to ship spent fuel to a central facility called ZWILAG for interim storage. In the mid-nineties, COGEMA LOGISTICS was contracted by KKL for the supply of the TN trademark a52L and TN trademark 97L transport and storage casks for BWR fuel types. In 2003, KKL also ordered from COGEMA LOGISTICS the supply of six TNae24 BHL transport and storage casks. This paper shows how all the three cask designs have responded to the KKL needs to ship and store BWR spent fuel. In addition, it highlights the already significant operational feedback of the TN trademark 52L and TN trademark 97L casks by the KKL and ZWILAG operators

  7. Transposon Tn5 encodes streptomycin resistance in nonenteric bacteria.

    OpenAIRE

    O'Neill, E A; Kiely, G M; Bender, R A

    1984-01-01

    Strains of Caulobacter crescentus, Pseudomonas putida, Acinetobacter calcoaceticus, Rhizobium meliloti, and Rhodopseudomonas sphaeroides carrying the kanamycin resistance-encoding transposon Tn5 were 15 to 500 times more resistant to streptomycin than transposon-free strains. The streptomycin resistance determinant, which is separable from the kanamycin resistance determinant of Tn5, was not expressed in Escherichia coli or Klebsiella aerogenes.

  8. Modular evolution of TnGBSs, a new family of integrative and conjugative elements associating insertion sequence transposition, plasmid replication, and conjugation for their spreading.

    Science.gov (United States)

    Guérillot, Romain; Da Cunha, Violette; Sauvage, Elisabeth; Bouchier, Christiane; Glaser, Philippe

    2013-05-01

    Integrative and conjugative elements (ICEs) have a major impact on gene flow and genome dynamics in bacteria. The ICEs TnGBS1 and TnGBS2, first identified in Streptococcus agalactiae, use a DDE transposase, unlike most characterized ICEs, which depend on a phage-like integrase for their mobility. Here we identified 56 additional TnGBS-related ICEs by systematic genome analysis. Interestingly, all except one are inserted in streptococcal genomes. Sequence comparison of the proteins conserved among these ICEs defined two subtypes related to TnGBS1 or TnGBS2. We showed that both types encode different conjugation modules: a type IV secretion system, a VirD4 coupling protein, and a relaxase and its cognate oriT site, shared with distinct lineages of conjugative elements of Firmicutes. Phylogenetic analysis suggested that TnGBSs evolved from two conjugative elements of different origins by the successive recruitment of a transposition module derived from insertion sequences (ISs). Furthermore, TnGBSs share replication modules with different plasmids. Mutational analyses and conjugation experiments showed that TnGBS1 and TnGBS2 combine replication and transposition upstream promoters for their transfer and stabilization. Despite an evolutionarily successful horizontal dissemination within the genus Streptococcus, these ICEs have a restricted host range. However, we reveal that for TnGBS1 and TnGBS2, this host restriction is not due to a transfer incompatibility linked to the conjugation machineries but most likely to their ability for transient maintenance through replication after their transfer.

  9. Circulating proteins in response to combined-modality therapy in rectal cancer identified by antibody array screening.

    Science.gov (United States)

    Kalanxhi, Erta; Hektoen, Helga Helseth; Meltzer, Sebastian; Dueland, Svein; Flatmark, Kjersti; Ree, Anne Hansen

    2016-07-26

    The increasingly complex programs of contemporary cancer therapy emphasize the need for biological indicators of both therapeutic response and adverse effects. One example is combined-modality treatment aimed at improving long-term outcome in patients with locally advanced rectal cancer, which commonly comes at the price of extended limits of patient tolerance. In a prospective study with intensified neoadjuvant treatment of rectal cancer patients, using an antibody array, the profiling of approximately 500 proteins was performed in serial serum samples collected at different stages of the treatment course. The small number of proteins whose levels significantly changed after induction neoadjuvant chemotherapy (NACT) expanded substantially following the sequential chemoradiotherapy (CRT) and persisted four weeks later at treatment evaluation before pelvic surgery. Serum levels of proteins selected for validation of the experimental design, lipocalin-2 and matrix metalloproteinase-9, declined after NACT and gradually reverted to baseline values during the remaining neoadjuvant course. Of note, the greater the decline in post-NACT and post-CRT matrix metalloproteinase-9 levels, the more favorable progression-free survival. No correlation was found, however, with diarrhea scores, the clinical correlate of adverse therapeutic effects. Even though the findings were indicative of only tumor and not normal tissue effects, multiplex immunoassay analysis of circulating proteins in patients undergoing combined-modality therapy may in principle dissect the contribution of the individual modalities to overall systemic responses in patient outcome and tolerance. ClinicalTrials.gov NCT00278694 ; registration date: January 16, 2006, retrospective to enrollment of the first 10 patients of the current report.

  10. Development of gastric cancer in nonatrophic stomach with highly active inflammation identified by serum levels of pepsinogen and Helicobacter pylori antibody together with endoscopic rugal hyperplastic gastritis.

    Science.gov (United States)

    Watanabe, Mika; Kato, Jun; Inoue, Izumi; Yoshimura, Noriko; Yoshida, Takeichi; Mukoubayashi, Chizu; Deguchi, Hisanobu; Enomoto, Shotaro; Ueda, Kazuki; Maekita, Takao; Iguchi, Mikitaka; Tamai, Hideyuki; Utsunomiya, Hirotoshi; Yamamichi, Nobutake; Fujishiro, Mitsuhiro; Iwane, Masataka; Tekeshita, Tatsuya; Mohara, Osamu; Ushijima, Toshikazu; Ichinose, Masao

    2012-12-01

    This study aimed to elucidate groups at high risk of developing cancer among patients with serologically identified Helicobacter pylori infection and nonatrophic stomach. Annual endoscopy was performed for a mean of 5.4 years in 496 asymptomatic middle-aged men who were H. pylori antibody-positive and pepsinogen (PG) test-negative. Subjects were stratified according to the activity of H. pylori-associated gastritis measured by serum levels of PG and H. pylori antibody, and/or by endoscopic findings of rugal hyperplastic gastritis (RHG), and cancer development was investigated. During the study period, seven cases of cancer developed in the cohort (incidence rate, 261/100,000 person-years), with 85.7% developing in the group showing a PGI/II ratio ≤ 3.0, reflecting active inflammation-based high PGII levels. Cancer incidence was significantly higher in this group (750/100,000 person-years) than in groups with less active gastritis. Furthermore, cancer incidence for this group was significantly higher in the subgroup with high H. pylori antibody titers than in the low-titer subgroup. Meanwhile, endoscopic findings revealed that 11.7% of subjects showed RHG reflecting localized highly active inflammation, and cancer risk was significantly higher in patients with RHG than in patients without. Combining the two serum tests and endoscopic examination for RHG allowed identification of subjects with more active gastritis and higher cancer risk. No cancer development was observed in these high-risk subjects after H. pylori eradication. Subjects with highly active gastritis identified by the two serological tests and endoscopic RHG constitute a group at high risk of cancer development with H. pylori-infected nonatrophic stomach. Copyright © 2012 UICC.

  11. Antigenic Fingerprinting following Primary RSV Infection in Young Children Identifies Novel Antigenic Sites and Reveals Unlinked Evolution of Human Antibody Repertoires to Fusion and Attachment Glycoproteins.

    Directory of Open Access Journals (Sweden)

    Sandra Fuentes

    2016-04-01

    Full Text Available Respiratory Syncytial Virus (RSV is the major cause of pneumonia among infants. Here we elucidated the antibody repertoire following primary RSV infection and traced its evolution through adolescence and adulthood. Whole genome-fragment phage display libraries (GFPDL expressing linear and conformational epitopes in the RSV fusion protein (F and attachment protein (G were used for unbiased epitope profiling of infant sera prior to and following RSV infection. F-GFPDL analyses demonstrated modest changes in the anti-F epitope repertoires post-RSV infection, while G-GFPDL analyses revealed 100-fold increase in number of bound phages. The G-reactive epitopes spanned the N- and C-terminus of the G ectodomain, along with increased reactivity to the central conserved domain (CCD. Panels of F and G antigenic sites were synthesized to evaluate sera from young children (<2 yr, adolescents (14-18 yr and adults (30-45 yr in SPR real-time kinetics assays. A steady increase in RSV-F epitope repertoires from young children to adults was observed using peptides and F proteins. Importantly, several novel epitopes were identified in pre-fusion F and an immunodominant epitope in the F-p27. In all age groups, antibody binding to pre-fusion F was 2-3 folds higher than to post-fusion form. For RSV-G, antibody responses were high following early RSV infection in children, but declined significantly in adults, using either G proteins or peptides. This study identified unlinked evolution of anti-F and anti G responses and supportive evidence for immune pressure driven evolution of RSV-G. These findings could help development of effective countermeasures including vaccines.

  12. Distinguishing Truncated and Normal MUC1 Glycoform Targeting from Tn-MUC1-Specific CAR T Cells

    DEFF Research Database (Denmark)

    Posey, Avery D; Clausen, Henrik; June, Carl H

    2016-01-01

    Genetically modified T cells expressing chimeric antigen receptors (CARs) demonstrate potent clinical antitumor effects in a variety of blood cancers. However, clinical activity in solid tumors has been disappointing and toxicity has been a serious concern (Lamers et al., 2013; Morgan et al., 2010......). We recently found that a CAR composed of a scFv antibody fragment specific for the Tn-glycoform of MUC1 had potent activity in preclinical models of blood cancer and adenocarcinoma (Posey et al., 2016)....

  13. BWR-spent fuel transport and storage with the TN trademark 9/4 and TN trademark 24BH casks

    International Nuclear Information System (INIS)

    Wattez, L.; Marguerat, Y.; Hoesli, C.

    2004-01-01

    The Swiss Nuclear Utilities have started in 2001 to store spent fuel in dry metallic dual-purpose casks in ZWILAG, the Swiss interim storage facility. BKW FMB Energy Ltd., as Muehleberg Nuclear Power Plant owner, is involved in this process and has selected to store its spent fuel, a new high capacity dual-purpose cask, the TN trademark 24BH. For the transport in a medium size cask, COGEMA LOGISTICS has developed a new cask, the TN trademark 9/4, to replace the NTL9 cask, which performed numerous transports of BWR spent fuel in the past decades. Licensed IAEA 1996, the TN trademark 9/4 is a 40 ton transport cask, for 7 BWR high burn-up spent fuel assemblies. The spent fuel assemblies can be transferred in the ZWILAG hot cell in the TN trademark 24BH cask. The first use of these casks took place in 2003. Ten TN trademark 9/4 transports were performed, and one TN trademark 24BH was loaded. After a brief presentation of the operational aspects, the paper will focus on the TN trademark 24BH high capacity dual purpose cask, the TN trademark 9/4 transport cask and describe in detail their characteristics and possibilities

  14. Upregulation of Mrps18a in breast cancer identified by selecting phage antibody libraries on breast tissue sections

    DEFF Research Database (Denmark)

    Sørensen, Karen Marie Juul; Meldgaard, Theresa; Melchjorsen, Connie Jenning

    2017-01-01

    . RESULTS: We identified the mitochondrial ribosomal protein s18a (Mrps18a) as a protein which is upregulated in breast cancer. However, Mrps18a was not homogeneously upregulated in all cancer cells, suggesting the existence of sub-populations within the tumor. The upregulation was not confined...... to cytokeratin 19 and cytokeratin 14 double positive cells. CONCLUSION: This study illustrates how phage display can be applied towards the discovery of proteins which exhibit changes in their expression patterns. We identified the mitochondrial protein Mrps18a as being upregulated in human breast cancer cells...

  15. 78 FR 7993 - Amendment of Class D and E Airspace; Tri-Cities, TN; Revocation of Class E Airspace; Tri-City, TN

    Science.gov (United States)

    2013-02-05

    ...). The SNPRM would list Hawkins County Airport and Virginia Highlands Airport under their own designator... feet above the surface for Hawkins County Airport, Rogersville, TN, and Virginia Highlands Airport, VA... TN E5 Rogersville, TN [New] Hawkins County Airport, TN (Lat. 36[deg]27'27'' N., long. 82[deg]53'06...

  16. Development of a candidate secondary reference procedure (immunoassay based measurement procedure of higher metrological order) for cardiac troponin I: I. Antibody characterization and preliminary validation.

    Science.gov (United States)

    Noble, James E; Bunk, David M; Christenson, Robert H; Cole, Kenneth D; He, Hua-Jun; Katrukha, Alexei G; Panteghini, Mauro; Porter, Robert A; Schimmel, Heinz; Tate, Jillian R; Wang, Lili

    2010-11-01

    In this study, the first steps in the development of a secondary reference measurement procedure (RMP) 'higher metrological order measurement procedure' to support the cardiac troponin I (cTnI) standardization initiative is described. The RMP should be used to assign values to serum-based secondary reference materials (RMs) without analytical artifacts causing bias. A multiplexed bead-based assay and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were used to identify the optimum monoclonal antibody pair (clones 560 and 19C7) for the RMP. Using these antibodies, an ELISA-based procedure was developed to accurately measure the main cTnI forms present in blood. The proposed RMP appears to show no bias when tested on samples containing various troponin complexes, phosphorylated and dephosphorylated forms, and heparin. The candidate assay displayed suitable linearity and sensitivity (limit of detection, 0.052 μg/L) for the measurement of the proposed cTnI secondary RMs. Preliminary comparison data on patient samples with a commercial cTnI assay are also provided to support the suitability of RMP for value assignment to RMs. Full validation and final assessment of the RMP will be performed through transferability and inter-comparison studies.

  17. Antibody Epitopes Identified in Critical Regions of Dengue Virus Nonstructural 1 Protein in Mouse Vaccination and Natural Human Infections.

    Science.gov (United States)

    Hertz, Tomer; Beatty, P Robert; MacMillen, Zachary; Killingbeck, Sarah S; Wang, Chunling; Harris, Eva

    2017-05-15

    Dengue is a global public health problem and is caused by four dengue virus (DENV) serotypes (DENV1-4). A major challenge in dengue vaccine development is that cross-reactive anti-DENV Abs can be protective or potentially increase disease via Ab-dependent enhancement. DENV nonstructural protein 1 (NS1) has long been considered a vaccine candidate as it avoids Ab-dependent enhancement. In this study, we evaluated survival to challenge in a lethal DENV vascular leak model in mice immunized with NS1 combined with aluminum and magnesium hydroxide, monophosphoryl lipid A + AddaVax, or Sigma adjuvant system+CpG DNA, compared with mice infected with a sublethal dose of DENV2 and mice immunized with OVA (negative control). We characterized Ab responses to DENV1, 2, and 3 NS1 using an Ag microarray tiled with 20-mer peptides overlapping by 15 aa and identified five regions of DENV NS1 with significant levels of Ab reactivity in the NS1 + monophosphoryl lipid A + AddaVax group. Additionally, we profiled the Ab responses to NS1 of humans naturally infected with DENV2 or DENV3 in serum samples from Nicaragua collected at acute, convalescent, and 12-mo timepoints. One region in the wing domain of NS1 was immunodominant in both mouse vaccination and human infection studies, and two regions were identified only in NS1-immunized mice; thus, vaccination can generate Abs to regions that are not targeted in natural infection and could provide additional protection against lethal DENV infection. Overall, we identified a small number of immunodominant regions, which were in functionally important locations on the DENV NS1 protein and are potential correlates of protection. Copyright © 2017 by The American Association of Immunologists, Inc.

  18. An Enterobacter plasmid as a new genetic background for the transposon Tn1331

    Directory of Open Access Journals (Sweden)

    Alavi MR

    2011-11-01

    Full Text Available Mohammad R Alavi1,2, Vlado Antonic2, Adrien Ravizee1, Peter J Weina3, Mina Izadjoo1,2, Alexander Stojadinovic21Division of Wound Biology and Translational Research, Armed Forces Institute of Pathology and American Registry of Pathology, Washington DC, 2Combat Wound Initiative Program, Walter Reed Army Medical Center, Washington DC, 3The Walter Reed Army Institute of Research, Silver Spring, MD, USABackground: Genus Enterobacter includes important opportunistic nosocomial pathogens that could infect complex wounds. The presence of antibiotic resistance genes in these microorganisms represents a challenging clinical problem in the treatment of these wounds. In the authors’ screening of antibiotic-resistant bacteria from complex wounds, an Enterobacter species was isolated that harbors antibiotic-resistant plasmids conferring resistance to Escherichia coli. The aim of this study was to identify the resistance genes carried by one of these plasmids.Methods: The plasmids from the Enterobacter isolate were propagated in E. coli and one of the plasmids, designated as pR23, was sequenced by the Sanger method using fluorescent dye-terminator chemistry on a genetic analyzer. The assembled sequence was annotated by search of the GenBank database.Results: Plasmid pR23 is composed of the transposon Tn1331 and a backbone plasmid that is identical to the plasmid pPIGDM1 from Enterobacter agglomerans. The multidrug-resistance transposon Tn1331, which confers resistance to aminoglycoside and beta lactam antibiotics, has been previously isolated only from Klebsiella. The Enterobacter plasmid pPIGDM1, which carries a ColE1-like origin of replication and has no apparent selective marker, appears to provide a backbone for propagation of Tn1331 in Enterobacter. The recognition sequence of Tn1331 transposase for insertion into pPIGDM1 is the pentanucleotide TATTA, which occurs only once throughout the length of this plasmid.Conclusion: Transposition of Tn1331 into

  19. EnviroAtlas - Memphis, TN - EnviroAtlas Community Boundary

    Data.gov (United States)

    U.S. Environmental Protection Agency — This EnviroAtlas dataset shows the boundary of the Memphis, TN EnviroAtlas Community. It represents the outside edge of all the block groups included in the...

  20. 76 FR 14855 - Television Broadcasting Services; Nashville, TN

    Science.gov (United States)

    2011-03-18

    ... FEDERAL COMMUNICATIONS COMMISSION 47 CFR Part 73 [MB Docket No. 11-29, RM-11622; DA 11-335] Television Broadcasting Services; Nashville, TN AGENCY: Federal Communications Commission. ACTION: Proposed... 73 Television, Television broadcasting. Federal Communications Commission. Kevin R. Harding...

  1. Host-dependent transposon Tn5-mediated streptomycin resistance.

    OpenAIRE

    De Vos, G F; Finan, T M; Signer, E R; Walker, G C

    1984-01-01

    Transposon Tn5 encodes streptomycin resistance in addition to kanamycin-neomycin resistance. This resistance was not detectable in Escherichia coli but was efficiently expressed in Rhizobium meliloti and certain other strains. By analysis of cloned Tn5 restriction endonuclease fragments, the streptomycin resistance (str) gene was located in the right-hand side of the central region as the transposon is conventionally drawn. Transcription of str appeared to originate at pL, the promoter for th...

  2. [3H]Azidodantrolene photoaffinity labeling, synthetic domain peptides and monoclonal antibody reactivity identify the dantrolene binding sequence on RyR1

    Energy Technology Data Exchange (ETDEWEB)

    Paul-Pletzer, Kalanethee; Yamamoto, Takeshi; Bhat, Manju B.; Ma, Jianjie; Ikemoto, Noriaki; Jimenez, Leslie S.; Morimoto, Hiromi; Williams, Philip G.; Parness, Jerome

    2002-06-14

    Dantrolene is a drug that suppresses intracellular Ca2+ release from sarcoplasmic reticulum in normal skeletal muscle and is used as a therapeutic agent in individuals susceptible to malignant hyperthermia. Though its precise mechanism of action has not been elucidated, we have identified the N-terminal region (amino acids 1-1400) of the skeletal muscle isoform of the ryanodine receptor (RyR1), the primary Ca2+ release channel in sarcoplasmic reticulum, as a molecular target for dantrolene using the photoaffinity analog [3H]azidodantrolene(1). Here, we demonstrate that heterologously expressed RyR1 retains its capacity to be specifically labeled with [3H]azidodantrolene,indicating that muscle specific factors are not required for this ligand-receptor interaction. Synthetic domain peptides of RyR1, previously shown to affect RyR1 function in vitro and in vivo, were exploited as potential drug binding site mimics and used in photoaffinity labeling experiments. Only DP1 and DP1-2, peptide s containing the amino acid sequence corresponding to RyR1 residues 590-609, were specifically labeled by [3H]azidodantrolene. A monoclonal anti-RyR1 antibody which recognizes RyR1 and its 1400 amino acid N-terminal fragment, recognizes DP1 and DP1-2 in both Western blots and immunoprecipitation assays, and specifically inhibits [3H]azidodantrolene photolabeling of RyR1 and its N-terminal fragment in sarcoplasmic reticulum. Our results indicate that synthetic domain peptides can mimic a native, ligand binding conformation in vitro, and that the dantrolene binding site and the epitope for the monoclonal antibody on RyR1 are equivalent and composed of amino-acids 590-609.

  3. Human antibody responses of patients living in endemic areas for schistosomiasis to the tegumental protein Sm29 identified through genomic studies.

    Science.gov (United States)

    Cardoso, F C; Pacífico, R N A; Mortara, R A; Oliveira, S C

    2006-06-01

    Surface proteins of schistosomes are exposed to host tissues and thus present as potential candidate molecules for the development of new intervention strategies. Herein, we have identified a new tegumental protein of Schistosoma mansoni, termed Sm29. In silico analysis revealed a signal peptide, three glycosylation sites and a transmembrane region on Sm29 amino acid sequence. Sm29 transcription in mammalian developmental stages cDNA libraries of S. mansoni was verified by PCR using specific primers for Sm29 nucleotide sequence and it revealed the presence of transcripts in schistosomula and adult worm stages of the parasite. Sm29 (40-169) fragment was produced in Escherichia coli and purified by affinity chromatography to be used in the immunological assays. Confocal microscopy confirmed bioinformatic studies, revealing that Sm29 is a membrane-bound protein localized on the tegument of S. mansoni adult worm. ELISA was performed using rSm29 protein to investigate the antibody isotype profile to Sm29 in sera of patients living in endemic areas for schistosomiasis. IgG1 and IgG3 subclass antibodies to rSm29 were predominant in sera of individuals naturally resistant to infection and resistant to re-infection whereas low levels of IgM, IgA or IgE were measured. Since, IgG1 and IgG3 are involved in parasite killing and in protective immunity the findings reported here suggest the use of Sm29 as a potential candidate vaccine against schistosomiasis.

  4. Failure of anti-T-cell receptor V beta antibodies to consistently identify a malignant T-cell clone in Sézary syndrome.

    Science.gov (United States)

    Bigler, R D; Boselli, C M; Foley, B; Vonderheid, E C

    1996-11-01

    Monoclonal antibodies (MAbs) reacting with the human T cell receptor (TCR) V beta or V alpha region have been shown to be almost as specific as a private idiotypic MAb in identifying T cell clones. When available, V beta-specific MAbs offer the ease of immunofluorescence analysis to identify and quantitate expanded malignant or nonmalignant T cell populations without requiring polymerase chain reaction (PCR) technology to evaluate expression of V beta gene families. The V beta expression of peripheral blood lymphocytes from twenty-three consecutive patients with Sézary syndrome has been analyzed by reverse transcriptase (RT)-PCR. Ten patients had malignant T cell clones that expressed a TCR V beta corresponding to a commercially available anti-V beta antibody. Immunofluorescence staining with anti-V beta MAbs showed a direct correlation with RT-PCR results in seven of ten patients. No false positive reactivity was noted on immunofluorescence staining with any MAb. Cells from three patients, however, did not react with the corresponding anti-V beta MAb. These three cases expressed a TCR V beta from gene families containing a single member, ie, V beta 14, V beta 18, and V beta 20, yet MAbs reported to be specific for these regions failed to react with the T cell clone from these patients. Sequencing of the PCR product in these cases confirmed the RT-PCR results. Cells from two patients expressed a TCR using V beta 5.1-D beta 1.1 genes with different J-C segments. One patient's cells reacted with an anti-V beta 5.1 MAb (LC4) whereas the other patient's cells bound one-tenth the amount of this same MAb. These results indicate that currently available anti-TCR V region MAbs may not react consistently with T cell clones expressing the corresponding V region or may react with a low affinity making detection difficult. Differences in the J-C junction or in CDR3 may influence the binding of these MAbs. Until the false negative rate is reduced and the fine specificity and

  5. Fully automated high-throughput chromatin immunoprecipitation for ChIP-seq: identifying ChIP-quality p300 monoclonal antibodies.

    Science.gov (United States)

    Gasper, William C; Marinov, Georgi K; Pauli-Behn, Florencia; Scott, Max T; Newberry, Kimberly; DeSalvo, Gilberto; Ou, Susan; Myers, Richard M; Vielmetter, Jost; Wold, Barbara J

    2014-06-12

    Chromatin immunoprecipitation coupled with DNA sequencing (ChIP-seq) is the major contemporary method for mapping in vivo protein-DNA interactions in the genome. It identifies sites of transcription factor, cofactor and RNA polymerase occupancy, as well as the distribution of histone marks. Consortia such as the ENCyclopedia Of DNA Elements (ENCODE) have produced large datasets using manual protocols. However, future measurements of hundreds of additional factors in many cell types and physiological states call for higher throughput and consistency afforded by automation. Such automation advances, when provided by multiuser facilities, could also improve the quality and efficiency of individual small-scale projects. The immunoprecipitation process has become rate-limiting, and is a source of substantial variability when performed manually. Here we report a fully automated robotic ChIP (R-ChIP) pipeline that allows up to 96 reactions. A second bottleneck is the dearth of renewable ChIP-validated immune reagents, which do not yet exist for most mammalian transcription factors. We used R-ChIP to screen new mouse monoclonal antibodies raised against p300, a histone acetylase, well-known as a marker of active enhancers, for which ChIP-competent monoclonal reagents have been lacking. We identified, validated for ChIP-seq, and made publicly available a monoclonal reagent called ENCITp300-1.

  6. Tn6350, a Novel Transposon Carrying Pyocin S8 Genes Encoding a Bacteriocin with Activity against Carbapenemase-Producing Pseudomonas aeruginosa.

    Science.gov (United States)

    Turano, Helena; Gomes, Fernando; Barros-Carvalho, Gesiele A; Lopes, Ralf; Cerdeira, Louise; Netto, Luis E S; Gales, Ana C; Lincopan, Nilton

    2017-05-01

    A novel transposon belonging to the Tn 3 -like family was identified on the chromosome of a commensal strain of Pseudomonas aeruginosa sequence type 2343 (ET02). Tn 6350 is 7,367 bp long and harbors eight open reading frames (ORFs), an ATPase (IS 481 family), a transposase (DDE catalytic type), a Tn 3 resolvase, three hypothetical proteins, and genes encoding the new pyocin S8 with its immunity protein. We show that pyocin S8 displays activity against carbapenemase-producing P. aeruginosa , including IMP-1, SPM-1, VIM-1, GES-5, and KPC-2 producers. Copyright © 2017 American Society for Microbiology.

  7. Highly efficient full-length hepatitis C virus genotype 1 (strain TN) infectious culture system

    DEFF Research Database (Denmark)

    Li, Yi-Ping; Ramirez, Santseharay; Jensen, Sanne B

    2012-01-01

    Chronic infection with hepatitis C virus (HCV) is an important cause of end stage liver disease worldwide. In the United States, most HCV-related disease is associated with genotype 1 infection, which remains difficult to treat. Drug and vaccine development was hampered by inability to culture...... with 8 changes (designated TN cell-culture derived, TNcc) replicated efficiently and released infectious particles of ∼5 log(10) focus-forming units per mL; passaged TNcc did not require additional changes. IFN-α and directly acting antivirals targeting the HCV protease, NS5A, and NS5B, each inhibited...... full-length TN infection dose-dependently. Given the unique importance of genotype 1 for pathogenesis, this infectious 1a culture system represents an important advance in HCV research. The approach used and the mutations identified might permit culture development for other HCV isolates, thus...

  8. Genetic analysis of attTn7, the transposon Tn7 attachment site in Escherichia coli, using a novel M13-based transduction assay.

    Science.gov (United States)

    Qadri, M I; Flores, C C; Davis, A J; Lichtenstein, C P

    1989-05-05

    The large (14 kb; kb = 10(3) bases) bacterial transposon, Tn7 (encoding resistance to trimethoprim and streptomycin/spectinomycin), has unusual properties. Like other elements, Tn7 transposes with low efficiency and low target-site specificity, but Tn7 also transposes, with high frequency in a unique orientation, to a preferred "attachment" site, called attTn7, in the Escherichia coli chromosome and similarly into plasmids containing attTn7. We developed a novel bacteriophage M13-based assay system to measure the transposition frequency of Tn7 to M13mp phage vectors containing attTn7 on a cloned 1 kb fragment of chromosomal DNA. Phage harvested from a Tn7 donor strain were used to infect recipient bacteria with selection for trimethoprim resistance. Transposition frequency, expressed as the number of trimethoprim-resistant colonies per plaque-forming unit, was found to be approximately 10(-4) to M13mp::attTn7, in contrast to 10(-10) to M13mp recombinants with approximately 1 kb insertions of other, "generic brand", DNA. By deletion analysis of M13mp::attTn7, we show that attTn7 is contained within a 64 base-pair region; sequences adjacent to the actual insertion site and encoding the carboxy terminus of the glmS gene are required. This assay also provided evidence for transposition immunity conferred by the right end of Tn7.

  9. Tetracycline alters gene expression in Salmonella strains that harbor the Tn10 transposon.

    Science.gov (United States)

    Hüttener, M; Prieto, A; Aznar, S; Dietrich, M; Paytubi, S; Juárez, A

    2018-04-01

    In this report, we show that bacterial plasmids that harbor the Tn10 transposon (i.e., the IncHI1 plasmid R27) modify expression of different Salmonella regulons responding to the presence of tetracycline (Tc) in the medium. By using as a model the Tc-dependent upregulation of the ibpAB operon (which belongs to the heat shock regulon), we have identified Tn10-tetA (coding for a Tc efflux pump) and adjacent tetC sequences as required for ibpAB upregulation. Characterization of transcripts in the tetAC region showed that tetA transcription can continue into tetC sequences, generating a long 3'UTR sequence, which can protect transcripts from RNA processing, thus increasing the expression of TetA protein. In the presence of Tc, the DnaK and IbpA chaperones are overexpressed and translocated to the periplasm and to the membrane fraction respectively. DnaK targeting unfolded proteins is known to induce heat shock by avoiding RpoH proteolysis. We correlate expression levels of Tn10-encoded TetA protein with heat shock induction in Salmonella, likely because TetA activity compromises protein secretion. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

  10. Transposon Tn5393e carrying the aphA1-containing transposon Tn6023 upstream of strAB does not confer resistance to streptomycin.

    Science.gov (United States)

    Cain, Amy K; Hall, Ruth M

    2011-09-01

    The simplest form of transposon Tn5393 carries the strA and strB genes that confer resistance to streptomycin, in addition to its transposition determinants. Tn5393e, made up of Tn5393 and a second transposon, Tn6023, was found in an IncHI2 plasmid, pSRC125, recovered from a multiply antibiotic-resistant Salmonella enterica serovar Typhimurium isolate of bovine origin. Tn6023 is made up of the aphA1b gene flanked by two inversely oriented copies of insertion sequence (IS) IS26 and is flanked by an 8 bp duplication. It is related to several other transposons that carry aphA1b in fragments of differing length, also flanked by copies of IS26. Tn6023 is located in the tnpR gene of Tn5393, which lies upstream of the strA and strB genes, and the combined structure was designated Tn5393e. Although neither strA nor strB contain any mutations that would inactivate them, pSRC125 does not confer resistance to streptomycin, indicating that the strA and strB genes are not expressed. In Tn5393, strA and strB are transcribed from the tnpR promoter, and in Tn5393e neither this transcript nor the transcript from the aphA1b promoter in Tn6023 must reach strAB. Tn5393e was previously found in different locations in Corynebacteria, indicating that it can move and suggesting a wide distribution. The structures of several further variants of Tn5393 found in GenBank were analyzed and assigned variant designations Tn5393f-Tn5393i.

  11. Anti-citrullinated heat shock protein 90 antibodies identified in bronchoalveolar lavage fluid are a marker of lung-specific immune responses.

    Science.gov (United States)

    Harlow, Lisa; Gochuico, Bernadette R; Rosas, Ivan O; Doyle, Tracy J; Osorio, Juan C; Travers, Timothy S; Camacho, Carlos C; Oddis, Chester V; Ascherman, Dana P

    2014-11-01

    Previous work has demonstrated a correlation between serum anti-citrullinated HSP90 antibodies and rheumatoid arthritis-associated interstitial lung disease (RA-ILD). To further investigate this potential pathogenic relationship, we used ELISA-based techniques to assess anti-citrullinated HSP90 antibody profiles in bronchoalveolar lavage fluid (BALF) of patients with different stages of RA-ILD. 9/21 RA-derived BALF specimens demonstrated IgG and/or IgA antibodies targeting citrullinated HSP90 proteins/peptides, highlighting disease specific responses (with a predilection for RA-ILD) that did not occur in IPF patients (0/5) or healthy control subjects (0/5). Comparison of antibody profiles between BALF and matching serum specimens revealed various recognition patterns favoring predominant production of anti-citrullinated HSP90 antibodies within the lung microenvironment-further supporting the connection between this antibody specificity and parenchymal lung disease. Equally important, qualitative as well as quantitative differences in anti-citrullinated HSP90 profiles between BALF and serum indicate that the lung plays a direct role in shaping the immune repertoire of RA/RA-ILD. Published by Elsevier Inc.

  12. Low hepatitis C antibody screening rates among an insured population of Tennessean Baby Boomers.

    Directory of Open Access Journals (Sweden)

    James G Carlucci

    Full Text Available Chronic Hepatitis C Virus (HCV infection is common and can cause liver disease and death. Persons born from 1945 through 1965 ("Baby Boomers" have relatively high prevalence of chronic HCV infection, prompting recommendations that all Baby Boomers be screened for HCV. If chronic HCV is confirmed, evaluation for antiviral treatment should be performed. Direct-acting antivirals can cure more than 90% of people with chronic HCV. This sequence of services can be referred to as the HCV "cascade of cure" (CoC. The Tennessee (TN Department of Health (TDH and a health insurer with presence in TN aimed to determine the proportion of Baby Boomers who access HCV screening services and appropriately navigate the HCV CoC in TN.TDH surveillance data and insurance claim records were queried to identify the cohort of Baby Boomers eligible for HCV testing. Billing codes and pharmacy records from 2013 through 2015 were used to determine whether HCV screening and other HCV-related services were provided. The proportion of individuals accessing HCV screening and other steps along the HCV CoC was determined. Multivariable analyses were performed to identify factors associated with HCV screening and treatment.Among 501,388 insured Tennessean Baby Boomers, 7% were screened for HCV. Of the 40,019 who received any HCV-related service, 86% were screened with an HCV antibody test, 20% had a confirmatory HCV PCR, 9% were evaluated for treatment, and 4% were prescribed antivirals. Hispanics were more likely to be screened and treated for HCV than non-Hispanic whites. HCV screening was more likely to occur in the Nashville-Davidson region than in other regions of TN, but there were regional variations in HCV treatment.Many insured Tennessean Baby Boomers do not access HCV screening services, despite national recommendations. Demographic and regional differences in uptake along the HCV CoC should inform public health interventions aimed at mitigating the effects of chronic

  13. 76 FR 33656 - Television Broadcasting Services; Nashville, TN

    Science.gov (United States)

    2011-06-09

    ... FEDERAL COMMUNICATIONS COMMISSION 47 CFR Part 73 [MB Docket No. 11-29; RM-11622, DA 11-949] Television Broadcasting Services; Nashville, TN AGENCY: Federal Communications Commission. ACTION: Final rule... Part 73 Television. Federal Communications Commission. Barbara A. Kreisman, Chief, Video Division...

  14. Japanese version transport/storage packaging 'TN24'

    International Nuclear Information System (INIS)

    Kakunai, H.; Iida, T.; Tsuda, K.; Akamatsu, H.

    1993-01-01

    Since 1983, Kobe Steel has been engaged jointly with the French company Transnucleaire in the development of 'TN24', a dry-type transport and storage packaging for irradiated fuel elements. This report describes the packaging, which has been adapted for use in domestic power stations using BWRs on the basis of the results of this development. (J.P.N.)

  15. A novel transposon, Tn6306, mediates the spread of blaIMI in Enterobacteriaceae in hospitals

    Directory of Open Access Journals (Sweden)

    Fangfang Zhang

    2017-12-01

    Full Text Available The increasing incidence of carbapenem-resistant Enterobacteriaceae has become a challenge for clinical therapy. In our study, we analysed the molecular characteristics of imipenem-hydrolyzing β-lactamase (IMI in Enterobacteriaceae isolates. Two reported clinical isolates, the IMI-3-producing Raoultella ornithinolytica RJ46C and the IMI-2-producing Escherichia coli RJ18 were identified in our retrospective review of isolates collected from June 2010 to June 2013, both isolates were resistant to carbapenem but sensitive to expanded-spectrum cephalosporins. The blaIMI gene was located on different ∼170-kb plasmids in both isolates. The blaIMI-3 gene was carried by the plasmid pRJ46C, which was extracted from the transconjugant and identified to be a 166,620-bp conjugative IncFIIY plasmid that contained 193 open reading frames, including replication-, plasmid conjugal transfer-, partitioning-, and mobilization-associated structures. The blaIMI-3 gene was located on a 15-kb region with a completely inverted sequence relative to that of plasmid pGA45, two ISEcl1-like elements containing two 33-bp complete inverted repeats were in an inverted orientation on both sides of the 15-kb region. We identified this typical structure as a novel composite transposon named Tn6306, indicating the occurrence of transposition. In addition, the blaIMI-2-carrying pRJ18 was an IncFIB plasmid, and a similar ISEcl1-like element was identified in an inverted direction upstream of IMI-2 in pRJ18. The identification of blaIMI in R. ornithinolytica and E. coli highlights the diversity of spreading carbapenemases in Enterobacteriaceae between hospitals and the environment in China. The novel transposon Tn6306, and other insert sequences, may play important roles in blaIMI mobilization. Keywords: blaIMI in Enterobacteriaceae, Genetic environment, Plasmids, Novel transposon structure Tn6306, ISEcl1-like element

  16. Chimeric recombinant antibody fragments in cardiac troponin I immunoassay.

    Science.gov (United States)

    Hyytiä, Heidi; Heikkilä, Taina; Brockmann, Eeva-Christine; Kekki, Henna; Hedberg, Pirjo; Puolakanaho, Tarja; Lövgren, Timo; Pettersson, Kim

    2015-03-01

    To introduce a novel nanoparticle-based immunoassay for cardiac troponin I (cTnI) utilizing chimeric antibody fragments and to demonstrate that removal of antibody Fc-part and antibody chimerization decrease matrix related interferences. A sandwich-type immunoassay for cTnI based on recombinant chimeric (mouse variable/human constant) antigen binding (cFab) antibodies and intrinsically fluorescent nanoparticles was developed. To test whether using chimeric antibody fragments helps to avoid matrix related interferences, samples (n=39) with known amounts of triglycerides, bilirubin, rheumatoid factor (RF) or human anti-mouse antibodies (HAMAs) were measured with the novel assay, along with a previously published nanoparticle-based research assay with the same antibody epitopes. The limit of detection (LoD) was 3.30ng/L. Within-laboratory precision for 29ng/L and 2819ng/L cTnI were 13.7% and 15.9%, respectively. Regression analysis with Siemens ADVIA Centaur® yielded a slope (95% confidence intervals) of 0.18 (0.17-1.19) and a y-intercept of 1.94 (-1.28-3.91) ng/L. When compared to a previously published nanoparticle-based assay, the novel assay showed substantially reduced interference in the tested interference prone samples, 15.4 vs. 51.3%. A rheumatoid factor containing sample was decreased from 241ng/L to TnI and decreased matrix related interferences, thus resulting in a lower number of falsely elevated cTnI-values. Copyright © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  17. Use of flow cytometry to identify monoclonal antibodies that recognize conserved epitopes on orthologous leukocyte differentiation antigens in goats, llamas, and rabbits

    Czech Academy of Sciences Publication Activity Database

    Davis, W. C.; Drbal, Karel; El-Aziz, A.; Mosaad, A.E.; Elbagory, A.R.M.; TIbary, A.; Barrington, G.M.; Park, Y.H.; Hamilton, M.J.

    2007-01-01

    Roč. 119, 1-2 (2007), s. 123-130 ISSN 0165-2427 Institutional research plan: CEZ:AV0Z50520514 Keywords : flow cytometry * monoclonal antibodies * leukocyte s Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.957, year: 2007

  18. Florfenicol-Chloramphenicol Exporter Gene fexA Is Part of the Novel Transposon Tn558

    Science.gov (United States)

    Kehrenberg, Corinna; Schwarz, Stefan

    2005-01-01

    The florfenicol-chloramphenicol exporter gene fexA is part of the novel transposon Tn558 from Staphylococcus lentus. Similarities between Tn558 and Tn554 from Staphylococcus aureus included the arrangement of the transposase genes tnpA to -C and an att554-like target sequence. Circular forms of Tn558 were detected and suggest the functional activity of this transposon. PMID:15673776

  19. Mobility and generation of mosaic non-autonomous transposons by Tn3-derived inverted-repeat miniature elements (TIMEs).

    Science.gov (United States)

    Szuplewska, Magdalena; Ludwiczak, Marta; Lyzwa, Katarzyna; Czarnecki, Jakub; Bartosik, Dariusz

    2014-01-01

    Functional transposable elements (TEs) of several Pseudomonas spp. strains isolated from black shale ore of Lubin mine and from post-flotation tailings of Zelazny Most in Poland, were identified using a positive selection trap plasmid strategy. This approach led to the capture and characterization of (i) 13 insertion sequences from 5 IS families (IS3, IS5, ISL3, IS30 and IS1380), (ii) isoforms of two Tn3-family transposons--Tn5563a and Tn4662a (the latter contains a toxin-antitoxin system), as well as (iii) non-autonomous TEs of diverse structure, ranging in size from 262 to 3892 bp. The non-autonomous elements transposed into AT-rich DNA regions and generated 5- or 6-bp sequence duplications at the target site of transposition. Although these TEs lack a transposase gene, they contain homologous 38-bp-long terminal inverted repeat sequences (IRs), highly conserved in Tn5563a and many other Tn3-family transposons. The simplest elements of this type, designated TIMEs (Tn3 family-derived Inverted-repeat Miniature Elements) (262 bp), were identified within two natural plasmids (pZM1P1 and pLM8P2) of Pseudomonas spp. It was demonstrated that TIMEs are able to mobilize segments of plasmid DNA for transposition, which results in the generation of more complex non-autonomous elements, resembling IS-driven composite transposons in structure. Such transposon-like elements may contain different functional genetic modules in their core regions, including plasmid replication systems. Another non-autonomous element "captured" with a trap plasmid was a TIME derivative containing a predicted resolvase gene and a res site typical for many Tn3-family transposons. The identification of a portable site-specific recombination system is another intriguing example confirming the important role of non-autonomous TEs of the TIME family in shuffling genetic information in bacterial genomes. Transposition of such mosaic elements may have a significant impact on diversity and evolution, not

  20. Mobility and generation of mosaic non-autonomous transposons by Tn3-derived inverted-repeat miniature elements (TIMEs.

    Directory of Open Access Journals (Sweden)

    Magdalena Szuplewska

    Full Text Available Functional transposable elements (TEs of several Pseudomonas spp. strains isolated from black shale ore of Lubin mine and from post-flotation tailings of Zelazny Most in Poland, were identified using a positive selection trap plasmid strategy. This approach led to the capture and characterization of (i 13 insertion sequences from 5 IS families (IS3, IS5, ISL3, IS30 and IS1380, (ii isoforms of two Tn3-family transposons--Tn5563a and Tn4662a (the latter contains a toxin-antitoxin system, as well as (iii non-autonomous TEs of diverse structure, ranging in size from 262 to 3892 bp. The non-autonomous elements transposed into AT-rich DNA regions and generated 5- or 6-bp sequence duplications at the target site of transposition. Although these TEs lack a transposase gene, they contain homologous 38-bp-long terminal inverted repeat sequences (IRs, highly conserved in Tn5563a and many other Tn3-family transposons. The simplest elements of this type, designated TIMEs (Tn3 family-derived Inverted-repeat Miniature Elements (262 bp, were identified within two natural plasmids (pZM1P1 and pLM8P2 of Pseudomonas spp. It was demonstrated that TIMEs are able to mobilize segments of plasmid DNA for transposition, which results in the generation of more complex non-autonomous elements, resembling IS-driven composite transposons in structure. Such transposon-like elements may contain different functional genetic modules in their core regions, including plasmid replication systems. Another non-autonomous element "captured" with a trap plasmid was a TIME derivative containing a predicted resolvase gene and a res site typical for many Tn3-family transposons. The identification of a portable site-specific recombination system is another intriguing example confirming the important role of non-autonomous TEs of the TIME family in shuffling genetic information in bacterial genomes. Transposition of such mosaic elements may have a significant impact on diversity and

  1. Neuronal Antibody Biomarkers for Sydenham’s Chorea Identify a New Group of Children with Chronic Recurrent Episodic Acute Exacerbations of Tic and Obsessive Compulsive Symptoms Following a Streptococcal Infection

    Science.gov (United States)

    Singer, Harvey S.; Mascaro-Blanco, Adda; Alvarez, Kathy; Morris-Berry, Christina; Kawikova, Ivana; Ben-Pazi, Hilla; Thompson, Carol B.; Ali, Syed F.; Kaplan, Edward L.; Cunningham, Madeleine W.

    2015-01-01

    Several autoantibodies (anti-dopamine 1 (D1R) and 2 (D2R) receptors, anti-tubulin, anti-lysoganglioside-GM1) and antibody-mediated activation of calcium calmodulin dependent protein kinase II (CaMKII) signaling activity are elevated in children with Sydenham’s chorea (SC). Recognizing proposed clinical and autoimmune similarities between SC and PANDAS (pediatric autoimmune neuropsychiatric disorder associated with a streptococcal infection), we sought to identify serial biomarker changes in a slightly different population. Antineuronal antibodies were measured in eight children (mean 11.3 years) with chronic, dramatic, recurrent tics and obsessive-compulsive disorder (OCD) associated with a group A β-hemolytic streptococcal (GABHS) respiratory tract infection, but differing because they lacked choreiform movements. Longitudinal serum samples in most subjects included two pre-exacerbation samples, Exac), one midst Exac (abrupt recurrence of tic/OCD; temporally association with a GABHS infection in six of eight subjects), and two post-Exac. Controls included four groups of unaffected children (n = 70; mean 10.8 years) obtained at four different institutions and published controls. Clinical exacerbations were not associated with a significant rise in antineuronal antibody titers. CaMKII activation was increased at the GABHS exacerbation point in 5/6 subjects, exceeded combined and published control’s 95th percentile at least once in 7/8 subjects, and median values were elevated at each time point. Anti-tubulin and anti-D2R titers did not differ from published or combined control group’s 95th percentile or median values. Differences in anti-lysoganglioside-GM1 and anti-D1R titers were dependent on the selected control. Variances in antibody titers and CaMKII activation were identified among the institutional control groups. Based on comparisons to published studies, results identify two groups of PANDAS: 1) a cohort, represented by this study, which lacks

  2. Lightcurve Analysis of Near-Earth Asteroid 2010 TN54

    Science.gov (United States)

    Warner, Brian D.; Benishek, Vladimir; Ferrero, Andrea

    2014-01-01

    CCD photometry observations of the near-Earth asteroid (NEA) 2010 TN 54 indicate a period of either 6.14 h or 12.12 h, depending on whether a monomodal or bimodal lightcurve is adopted. The amplitude was only 0.07 ± 0.01 mag, which - along with the period being nearly commensurate with an Earth day - made finding a definitive solution difficult, despite being observed from locations in North America and Europe.

  3. Multiple antibody targets on herpes B glycoproteins B and D identified by screening sera of infected rhesus macaques with peptide microarrays.

    Directory of Open Access Journals (Sweden)

    Sven-Kevin Hotop

    Full Text Available Herpes B virus (or Herpesvirus simiae or Macacine herpesvirus 1 is endemic in many populations of macaques, both in the wild and in captivity. The virus elicits only mild clinical symptoms (if any in monkeys, but can be transmitted by various routes, most commonly via bites, to humans where it causes viral encephalitis with a high mortality rate. Hence, herpes B constitutes a considerable occupational hazard for animal caretakers, veterinarians and laboratory personnel. Efforts are therefore being made to reduce the risk of zoonotic infection and to improve prognosis after accidental exposure. Among the measures envisaged are serological surveillance of monkey colonies and specific diagnosis of herpes B zoonosis against a background of antibodies recognizing the closely related human herpes simplex virus (HSV. 422 pentadecapeptides covering, in an overlapping fashion, the entire amino acid sequences of herpes B proteins gB and gD were synthesized and immobilized on glass slides. Antibodies present in monkey sera that bind to subsets of the peptide collection were detected by microserological techniques. With 42 different rhesus macaque sera, 114 individual responses to 18 different antibody target regions (ATRs were recorded, 17 of which had not been described earlier. This finding may pave the way for a peptide-based, herpes B specific serological diagnostic test.

  4. Characterization of Tn6238 with a New Allele of aac(6′)-Ib-cr

    Science.gov (United States)

    Quiroga, María P.; Orman, Betina; Errecalde, Laura; Kaufman, Sara

    2015-01-01

    Here, we report that the genetic structure of Tn1331 remained conserved in Argentina from 1989 to 2013 (72 of 73 isolates), with the exception being the plasmid-borne Tn1331-like transposon Tn6238 containing a new aac(6′)-Ib-cr allele recovered from a colistin-resistant Klebsiella pneumoniae clinical isolate. A bioinformatic analysis of aac(6′)-Ib-like gene cassettes suggests that this new aac(6′)-Ib-cr allele emerged through mutation or homologous recombination in the Tn1331 genetic platform. Tn6238 is a novel platform for the dissemination of aminoglycoside and fluoroquinolone resistance determinants. PMID:25691640

  5. Flow cytometry-based assay to evaluate human serum MUC1-Tn antibodies

    DEFF Research Database (Denmark)

    Van Elssen, Catharina H M J; Clausen, Henrik; Germeraad, Wilfred T V

    2011-01-01

    Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylat......Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due...

  6. Metabolite profiling reveals abiotic stress tolerance in Tn5 mutant of Pseudomonas putida.

    Directory of Open Access Journals (Sweden)

    Vasvi Chaudhry

    Full Text Available Pseudomonas is an efficient plant growth-promoting rhizobacteria (PGPR; however, intolerance to drought and high temperature limit its application in agriculture as a bioinoculant. Transposon 5 (Tn5 mutagenesis was used to generate a stress tolerant mutant from a PGPR Pseudomonas putida NBRI1108 isolated from chickpea rhizosphere. A mutant NBRI1108T, selected after screening of nearly 10,000 transconjugants, exhibited significant tolerance towards high temperature and drought. Southern hybridization analysis of EcoRI and XhoI restricted genomic DNA of NBRI1108T confirmed that it had a single Tn5 insertion. The metabolic changes in the polar and non-polar extracts of NBRI1108 and NBRI1108T were examined using 1H, 31P nuclear magnetic resonance (NMR spectroscopy and gas chromatography-mass spectrometry (GC-MS. Thirty six chemically diverse metabolites consisting of amino acids, fatty acids and phospholipids were identified and quantified. Insertion of Tn5 influenced amino acid and phospholipid metabolism and resulted in significantly higher concentration of aspartic acid, glutamic acid, glycinebetaine, glycerophosphatidylcholine (GPC and putrescine in NBRI1108T as compared to that in NBRI1108. The concentration of glutamic acid, glycinebetaine and GPC increased by 34%, 95% and 100%, respectively in the NBRI1108T as compared to that in NBRI1108. High concentration of glycerophosphatidylethanolamine (GPE and undetected GPC in NBRI1108 indicates that biosynthesis of GPE may have taken place via the methylation pathway of phospholipid biosynthesis. However, high GPC and low GPE concentration in NBRI1108T suggest that methylation pathway and phosphatidylcholine synthase (PCS pathway of phospholipid biosynthesis are being followed in the NBRI1108T. Application of multivariate principal component analysis (PCA on the quantified metabolites revealed clear variations in NBRI1108 and NBRI1108T in polar and non-polar metabolites. Identification of abiotic

  7. Interdisciplinary orthodontic treatment for a patient with generalized aggressive periodontitis: Assessment of IgG antibodies to identify type of periodontitis and correct timing of treatment.

    Science.gov (United States)

    Ishihara, Yoshihito; Tomikawa, Kazuya; Deguchi, Toru; Honjo, Tadashi; Suzuki, Koji; Kono, Takayuki; Kuboki, Takuo; Kamioka, Hiroshi; Takashiba, Shogo; Yamashiro, Takashi

    2015-06-01

    Aggressive periodontitis is a great challenge to clinicians when providing orthodontic treatment because of the potential for progression of periodontal disease. In this article, we report the successful comprehensive orthodontic treatment of bimaxillary protrusion and severe crowding in an adult with generalized aggressive periodontitis. A woman, aged 22 years 7 months, with a chief complaint of incisal crowding was diagnosed with a skeletal Class I malocclusion associated with severe anterior crowding, possibly worsened by generalized aggressive periodontitis. In addition to a periodontal examination, a blood IgG antibody titer analysis and microbiologic examination for periodontal pathogens were used to diagnose the type of periodontal disease and determine the proper timing to initiate orthodontic treatment. The total active treatment period was 28 months, followed by periodontal prostheses and regeneration therapy. Consequently, satisfactory facial profile, occlusion, and periodontal health were maintained for at least 36 months. These results indicate that efficient screening is important for providing successful orthodontic treatment in patients with advanced periodontal disease. This report also demonstrates the diagnostic importance of blood IgG antibody titer assays and microbiologic examinations to detect periodontal pathogens. Copyright © 2015 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.

  8. Antibody informatics for drug discovery

    DEFF Research Database (Denmark)

    Shirai, Hiroki; Prades, Catherine; Vita, Randi

    2014-01-01

    to the antibody science in every project in antibody drug discovery. Recent experimental technologies allow for the rapid generation of large-scale data on antibody sequences, affinity, potency, structures, and biological functions; this should accelerate drug discovery research. Therefore, a robust bioinformatic...... infrastructure for these large data sets has become necessary. In this article, we first identify and discuss the typical obstacles faced during the antibody drug discovery process. We then summarize the current status of three sub-fields of antibody informatics as follows: (i) recent progress in technologies...... for antibody rational design using computational approaches to affinity and stability improvement, as well as ab-initio and homology-based antibody modeling; (ii) resources for antibody sequences, structures, and immune epitopes and open drug discovery resources for development of antibody drugs; and (iii...

  9. [Identification of a high ammonia nitrogen tolerant and heterotrophic nitrification-aerobic denitrification bacterial strain TN-14 and its nitrogen removal capabilities].

    Science.gov (United States)

    Xin, Xin; Yao, Li; Lu, Lei; Leng, Lu; Zhou, Ying-Qin; Guo, Jun-Yuan

    2014-10-01

    A new strain of high ammonia nitrogen tolerant and heterotrophic nitrification-aerobic denitrification bacterium TN-14 was isolated from the environment. Its physiological and biochemical characteristics and molecular identification, performences of heterotrophic nitrification-aerobic, the abilities of resistance to ammonia nitrogen as well as the decontamination abilities were studied, respectively. It was preliminary identified as Acinetobacter sp. according to its physiological and biochemical characteristics and molecular identification results. In heterotrophic nitrification system, the ammonia nitrogen and total nitrogen removal rate of the bacterial strain TN-14 could reach 97.13% and 93.53% within 24 h. In nitrates denitrification system, the nitrate concentration could decline from 94.24 mg · L(-1) to 39.32 mg · L(-1) within 24 h, where the removal rate was 58.28% and the denitrification rate was 2.28 mg · (L · h)(-1); In nitrite denitrification systems, the initial concentration of nitrite could be declined from 97.78 mg · L(-1) to 21.30 mg x L(-1), with a nitrite nitrogen removal rate of 78.22%, and a denitrification rate of 2.55 mg · (L· h)(-1). Meanwhile, strain TN-14 had the capability of flocculant production, and the flocculating rate could reach 94.74% when its fermentation liquid was used to treat 0.4% kaolin suspension. Strain TN-14 could grow at an ammonia nitrogen concentration as high as 1200 mg · L(-1). In the aspect of actual piggery wastewater treatment by strain TN-14, the removal rate of COD, ammonia nitrogen, TN and TP cloud reached 85.30%, 65.72%, 64.86% and 79.41%, respectively. Strain TN-14 has a good application prospect in biological treatment of real high- ammonia wastewater.

  10. Preparation and Characterization of cis- and trans-[Ir(tn)2Cl2]CF3SO3 and of [Ir(tn)3]Cl3 (tn=propane-1,3-diamine)

    DEFF Research Database (Denmark)

    Brorson, Michael; Galsbøl, Frode; Simonsen, Kim

    1998-01-01

    Procedures are given for the preparation and isolation of cis- and trans-[Ir(tn)2Cl2]CF3SO3 and of [Ir(tn)3]Cl3, (tn=propane-1,3-diamine). The compounds were prepared by the use of Ir(thtp)3Cl3 (thtp=tetrahydrothiophene) as starting material, using either DMSO or neat tn as solvent. A procedure...... for the preparation of [Rh(tn)3]Cl3 in quantitative yield from Rh(thtp)3Cl3 is also given. The complexes were characterized by 1H and 13C NMR and by UV/VIS spectroscopy. The conformation of the six-membered chelate rings of [Ir(tn)3]3+ in the solid state was determined by single-crystal X-ray diffraction of [Ir(tn)3......] [Co(CN)6] x 5H2O. The three chelate rings all adopt the energetically favoured chair conformation; however, the overall idealized symmetry is C1. A comparative ligand field analysis, based on Gaussian resolution of the solution UV/VIS spectra for a number of homoleptic [M(N6)]3+ (M=CoIII, RhIII, Ir...

  11. Broadly neutralizing antibodies targeted to mucin-type carbohydrate epitopes of human immunodeficiency virus

    DEFF Research Database (Denmark)

    Hansen, J E; Nielsen, C; Arendrup, M

    1991-01-01

    The cancer-related mucin-type carbohydrate neoantigen Tn was found on gp160 and gp120 of human immunodeficiency virus (HIV). Immunoglobulin G (IgG) and IgM monoclonal antibodies (MAbs) against Tn neutralized infection with cell-free virus and blocked fusion between HIV-infected and uninfected cells....... This inhibition was found in infection of both lymphocytic cells and monocytoid cells. Viruses tested included six HIV-1 and five HIV-2 isolates propagated in different cells, as well as infectious plasma from AIDS patients. The antiviral effect of anti-Tn MAbs occurred by specific binding of the MAb to the virus...

  12. HTR-TN achievements and prospects for future developments

    International Nuclear Information System (INIS)

    Hittner, D.; Angulo, C.; Basini, V.; Bogusch, E.; Breuil, E.; Buckthorpe, D.; Chauvet, V.; Futterer, M.A.; Van Heek, A.; Von Lensa, W.; Yvon, P.

    2011-01-01

    It is already 10 years since the (European) High Temperature Reactor Technology Network (HTR-TN) launched a program for development of HTR technology, which expanded through three successive Euratom framework programs, with many projects in line with the network strategy. Widely relying in the beginning on the legacy of the former European HTR developments (DRAGON, AVR, THTR, etc.) that it contributed to safeguard, this program led to advances in HTR/VHTR technologies and produced significant results, which can contribute to the international cooperation through Euratom involvement in the Generation IV International Forum (GIF). the main achievements of the European program, performed in complement to efforts made in several European countries and other GIF partners, are presented: they concern the validation of computer codes (reactor physics, as well as system transient analysis from normal operation to air ingress accident and fuel performance in normal and accident conditions), materials (metallic materials for vessel, direct cycle turbines and intermediate heat exchanger, graphite, etc.), component development, fuel manufacturing and irradiation behavior, and specific HTR waste management (fuel and graphite). Key experiments have been performed or are still ongoing, like irradiation of graphite and of fuel material (PYCASSO experiment), high burn-up fuel PIE, safety test and isotopic analysis, IHX mock-up thermohydraulic test in helium atmosphere, air ingress experiment for a block type core, etc. Now HTR-TN partners consider that it is time for Europe to go a step forward toward industrial demonstration. In line with the orientations of the 'Strategic Energy Technology Plan (SET-Plan)' recently issued by the European Commission that promotes a strategy for development of low-carbon energy technologies and mentions Generation IV nuclear systems as part of key technologies, HTR-TN proposes to launch a program for extending the contribution of nuclear energy to

  13. Analisis Perubahan Kawasan Mangrove Berdasarkan Interpretasi Data Spasial Di Tn. Sembilang, Pantai Timur Sumatera, Banyuasin, Sumsel

    Directory of Open Access Journals (Sweden)

    Yetty Hastiana

    2012-02-01

    Full Text Available Due to the importance of mangrove ecosystem role to coastal area stability, study and research on mangrove ecosystem is interesting. Several study forms can be performed including by sightseeing and predicting degradation and change of mangrove conservation area during certain time. Result of prediction and analysis can be used by decision maker to state the priority of area protection. As intial step in management analysis for mangrove area ecosystem in Pasut area , TN. Sembilang Pantai Timur Sumatera, Banyuasin, SumSel,interpretation and identification can be performed during six years since it was stated as National park in 2003. Several techniques can be used for analyzing the ecosystem changes, one of these is by using remote sensing. In this research, remote sensing approach by landsat profile data from 2003 and 2009. The use of landsat data sequentially was aimed to interpret and identify changes in mangrove area during the time. Result of research showed that during six years there was changes and degradation mangrove ecosystem to be non mangrove of 14,57 %. This analysis hopely can be used as reference to apply wisdom and strategy of coastal area management . Analysis and strategic approach is become part of area optimation to reduce environmental pressures including biodiversity protection, coastal area protection also small islands from global climate change effect.Keywords: Spatial Analysis, Mangrove Ecosystem, Remote Sensing, TN. Sembilang SumSel.

  14. A role for Tn6029 in the evolution of the complex antibiotic resistance gene loci in genomic island 3 in enteroaggregative hemorrhagic Escherichia coli O104:H4.

    Directory of Open Access Journals (Sweden)

    Piklu Roy Chowdhury

    Full Text Available In enteroaggregative hemorrhagic Escherichia coli (EAHEC O104 the complex antibiotic resistance gene loci (CRL found in the region of divergence 1 (RD1 within E. coli genomic island 3 (GI3 contains blaTEM-1, strAB, sul2, tet(AA, and dfrA7 genes encoding resistance to ampicillin, streptomycin, sulfamethoxazole, tetracycline and trimethoprim respectively. The precise arrangement of antibiotic resistance genes and the role of mobile elements that drove the evolutionary events and created the CRL have not been investigated. We used a combination of bioinformatics and iterative BLASTn searches to determine the micro-evolutionary events that likely led to the formation of the CRL in GI3 using the closed genome sequences of EAHEC O104:H4 strains 2011C-3493 and 2009EL-2050 and high quality draft genomes of EAHEC E. coli O104:H4 isolates from sporadic cases not associated with the initial outbreak. Our analyses indicate that the CRL in GI3 evolved from a progenitor structure that contained an In2-derived class 1 integron in a Tn21/Tn1721 hybrid backbone. Within the hybrid backbone, a Tn6029-family transposon, identified here as Tn6029C abuts the sul1 gene in the 3'-Conserved Segment (-CS of a class 1 integron generating a unique molecular signature that has only previously been observed in pASL01a, a small plasmid found in commensal E. coli in West Africa. From this common progenitor, independent IS26-mediated events created two novel transposons identified here as Tn6029D and Tn6222 in 2011C-3493 and 2009EL-2050 respectively. Analysis of RD1 within GI3 reveals IS26 has played a crucial role in the assembly of regions within the CRL.

  15. Antibiotic and antiseptic resistance genes are linked on a novel mobile genetic element: Tn6087

    Science.gov (United States)

    Ciric, Lena; Mullany, Peter; Roberts, Adam P.

    2011-01-01

    Objectives Tn916-like elements are one of the most common types of integrative and conjugative element (ICE). In this study we aimed to determine whether novel accessory genes, i.e. genes whose products are not involved in mobility or regulation, were present on a Tn916-like element (Tn6087) isolated from Streptococcus oralis from the human oral cavity. Methods A minocycline-resistant isolate was analysed using restriction fragment length polymorphism (RFLP) analysis on amplicons derived from Tn916 and DNA sequencing to determine whether there were genetic differences in Tn6087 compared with Tn916. Mutational analysis was used to determine whether the novel accessory gene found was responsible for an observed extra phenotype. Results A novel Tn916-like element, Tn6087, is described that encodes both antibiotic and antiseptic resistance. The antiseptic resistance protein is encoded by a novel small multidrug resistance gene, designated qrg, that was shown to encode resistance to cetyltrimethylammonium bromide (CTAB), also known as cetrimide bromide. Conclusions This is the first Tn916-like element described that confers both antibiotic and antiseptic resistance, suggesting that selection of either antibiotic or antiseptic resistance will also select for the other and further highlights the need for prudent use of both types of compound. PMID:21816764

  16. Biodegradation of didecyldimethylammonium chloride by Pseudomonas fluorescens TN4 isolated from activated sludge.

    Science.gov (United States)

    Nishihara, T; Okamoto, T; Nishiyama, N

    2000-04-01

    Bacteria that degrade didecyldimethylammonium chloride (DDAC) were isolated from activated sludge from a municipal sewage treatment plant by enrichment culture with DDAC as a sole carbon source. One of the isolates, Pseudomonas fluorescens TN4, degraded DDAC to produce decyldimethylamine and subsequently, dimethylamine, as the intermediates. The TN4 strain also assimilated the other quaternary ammonium compounds (QACs), alkyltrimethyl- and alkylbenzyldimethyl-ammonium salts, but not alkylpyridinium salts. TN4 was highly resistant to these QACs and degraded them by an N-dealkylation process. These data mean that there are some QAC-resistant and QAC-degrading bacteria such as TN4 in the environment.

  17. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    surface expression of various antibody formats in the generated knockout strain. Functional scFv and scFab fragments were efficiently displayed on yeast whereas impaired chain assembly and heavy chain degradation was observed for display of full-length IgG molecules. To identify the optimal polypeptide......-antibody interface and the antibody intraface.the microenvironment and ecology of Acaryochloris and Prochloron, and in this thesis we attempted to further describe the distribution, growth characteristics and adaptive/regulatory mechanisms of these two cyanobacteria, both in their natural habitat and under defined...

  18. Mini-Tn7 Insertion in Bacteria With Multiple glmS-Linked attTn7 Sites: Example Burkholderia Mallei ATCC 23344

    National Research Council Canada - National Science Library

    Choi, Kyoung-Hee; DeShazer, David; Schweizer, Herbert P

    2006-01-01

    We recently constructed a series of broad-range mini-Tn7 vectors to address some of the problems inherent to plasmid-based cloning systems such as multi-copy state, necessity for continued selection...

  19. Mini-Tn7 Insertion in Bacteria With Multiple glmS-Linked attTn7 Sites: Example Burkholderia Mallei ATCC 23344

    National Research Council Canada - National Science Library

    Choi, Kyoung-Hee; DeShazer, David; Schweizer, Herbert P

    2006-01-01

    ... and scarcity for many bacteria. The potential of the mini-Tn7 vectors for finding wide-spread biomedical and environmental applications, particularly for analysis of complex systems such as animal and biofilm models was demonstrated...

  20. Antiprothrombin Antibodies

    Directory of Open Access Journals (Sweden)

    Polona Žigon

    2015-05-01

    Full Text Available In patients with the antiphospholipid syndrome (APS, the presence of a group of pathogenic autoantibodies called antiphospholipid antibodies causes thrombosis and pregnancy complications. The most frequent antigenic target of antiphospholipid antibodies are phospholipid bound β2-glycoprotein 1 (β2GPI and prothrombin. The international classification criteria for APS connect the occurrence of thrombosis and/or obstetric complications together with the persistence of lupus anticoagulant, anti-cardiolipin antibodies (aCL and antibodies against β2GPI (anti-β2GPI into APS. Current trends for the diagnostic evaluation of APS patients propose determination of multiple antiphospholipid antibodies, among them also anti-prothrombin antibodies, to gain a common score which estimates the risk for thrombosis in APS patients. Antiprothrombin antibodies are common in APS patients and are sometimes the only antiphospholipid antibodies being elevated. Methods for their determination differ and have not yet been standardized. Many novel studies confirmed method using phosphatidylserine/prothrombin (aPS/PT ELISA as an antigen on solid phase encompass higher diagnostic accuracy compared to method using prothrombin alone (aPT ELISA. Our research group developed an in-house aPS/PT ELISA with increased analytical sensitivity which enables the determination of all clinically relevant antiprothrombin antibodies. aPS/PT exhibited the highest percentage of lupus anticoagulant activity compared to aCL and anti-β2GPI. aPS/PT antibodies measured with the in-house method associated with venous thrombosis and presented the strongest independent risk factor for the presence of obstetric complications among all tested antiphospholipid antibodies

  1. Aberrantly glycosylated MUC1 is expressed on the surface of breast cancer cells and a target for antibody-dependent cell-mediated cytotoxicity

    DEFF Research Database (Denmark)

    Lavrsen, Kirstine; Madsen, Caroline B; Rasch, Morten G

    2013-01-01

    not covered by immunological tolerance in MUC1 humanized mice and man. The objective of this study was to determine if mouse antibodies to this Tn-MUC1 epitope induce antibody-dependent cellular cytotoxicity (ADCC) pivotal for their potential use in cancer immunotherapy. Binding affinity of mAb 5E5 directed...... to Tn-MUC1 was investigated using BiaCore. The availability of Tn-MUC1 on the surface of breast cancer cells was evaluated by immunohistochemistry, confocal microscopy, and flow cytometry, followed by in vitro assessment of antibody-dependent cellular cytotoxicity by mAb 5E5. Biacore analysis...... is expressed on the surface of breast cancer cells and a target for antibody-dependent cell-mediated cytotoxicity suggesting that antibodies targeting glycopeptide epitopes on mucins are strong candidates for cancer-specific immunotherapies....

  2. Base flipping in tn10 transposition: an active flip and capture mechanism.

    Directory of Open Access Journals (Sweden)

    Julien Bischerour

    2009-07-01

    Full Text Available The bacterial Tn5 and Tn10 transposases have a single active site that cuts both strands of DNA at their respective transposon ends. This is achieved using a hairpin intermediate that requires the DNA to change conformation during the reaction. In Tn5 these changes are controlled in part by a flipped nucleoside that is stacked on a tryptophan residue in a hydrophobic pocket of the transposase. Here we have investigated the base flipping mechanism in Tn10 transposition. As in Tn5 transposition, we find that base flipping takes place after the first nick and is required for efficient hairpin formation and resolution. Experiments with an abasic substrate show that the role of base flipping in hairpin formation is to remove the base from the DNA helix. Specific interactions between the flipped base and the stacking tryptophan residue are required for hairpin resolution later in the reaction. We show that base flipping in Tn10 transposition is not a passive reaction in which a spontaneously flipped base is captured and retained by the protein. Rather, it is driven in part by a methionine probe residue that helps to force the flipped base from the base stack. Overall, it appears that base flipping in Tn10 transposition is similar to that in Tn5 transposition.

  3. Human antibody responses of patients living in endemic areas for schistosomiasis to the tegumental protein Sm29 identified through genomic studies

    OpenAIRE

    Cardoso, F. C.; Pacifico, RNA; Mortara, Renato Arruda [UNIFESP; Oliveira, S. C.

    2006-01-01

    Surface proteins of schistosomes are exposed to host tissues and thus present as potential candidate molecules for the development of new intervention strategies. Herein, we have identified a new tegumental protein of Schistosoma mansoni, termed Sm29. in silico analysis revealed a signal peptide, three glycosylation sites and a transmembrane region on Sm29 amino acid sequence. Sm29 transcription in mammalian developmental stages cDNA libraries of S. mansoni was verified by PCR using specific ...

  4. Ultrasensitive cardiac troponin I antibody based nanohybrid sensor for rapid detection of human heart attack.

    Science.gov (United States)

    Bhatnagar, Deepika; Kaur, Inderpreet; Kumar, Ashok

    2017-02-01

    An ultrasensitive cardiac troponin I antibody conjugated with graphene quantum dots (GQD) and polyamidoamine (PAMAM) nanohybrid modified gold electrode based sensor was developed for the rapid detection of heart attack (myocardial infarction) in human. Screen printed gold (Au) electrode was decorated with 4-aminothiophenol for amine functionalization of the Au surface. These amino groups were further coupled with carboxyl functionalities of GQD with EDC-NHS reaction. In order to enhance the sensitivity of the sensor, PAMAM dendrimer was successively embedded on GQD through carbodiimide coupling to provide ultra-high surface area for antibody immobilization. The activated cardiac troponin I (cTnI) monoclonal antibody was immobilized on PAMAM to form nanoprobe for sensing specific heart attack marker cTnI. Various concentrations of cardiac marker, cTnI were electrochemically measured using cyclic voltammetry (CV) and differential pulse voltammetry (DPV) in human blood serum. The modifications on sensor surface were characterized by FTIR and AFM techniques. The sensor is highly specific to cTnI and showed negligible response to non-specific antigens. The sensitivity of the sensor was 109.23μAcm -2 μg -1 and lower limit of detection of cTnI was found 20fgmL -1 . Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Abrasive wear of Hilong BoTN hardfacings

    Science.gov (United States)

    Fedorova, L.; Fedorov, S.; Sadovnikov, A.; Ivanova, Y.; Voronina, M.

    2018-02-01

    The spread of steels, which are used to produce locks of steel drill pipes, adversely affects their wear resistance, which, in combination with low hardness of HV 2400 ... 2800 MPa as well as of the thread of screw, results in low wear resistance and the need for their reconstruction at the pipe control shop. An efficient way of improving the quality of drill pipe jonts is to hard-face them by the outside diameter with wear-resistant materials (hardbanding). One of the companies engaged in the development of hardfacing materials and hardbanding is Hilong (China) with weld seams of the brand BoTn. According to the results of the studies the following conclusion can be made: hardfacing increases the durability of the hardware, contributing to an increase in wear resistance of locks of DP under the conditions of abrasive action of aggressive geological formations; the usage of DP without wear-resistant weld seams is impermissible, because their further operation, as part of the drill-stem, can lead to emergency consequences; application of the pipes with the hardfacing collars together with the collars without hardfacing, due to varying degree of wear of jonts in the drill-stem, is also impermissible.

  6. Tn6188 - a novel transposon in Listeria monocytogenes responsible for tolerance to benzalkonium chloride.

    Directory of Open Access Journals (Sweden)

    Anneliese Müller

    Full Text Available Controlling the food-borne pathogen Listeria (L. monocytogenes is of great importance from a food safety perspective, and thus for human health. The consequences of failures in this regard have been exemplified by recent large listeriosis outbreaks in the USA and Europe. It is thus particularly notable that tolerance to quaternary ammonium compounds such as benzalkonium chloride (BC has been observed in many L. monocytogenes strains. However, the molecular determinants and mechanisms of BC tolerance of L. monocytogenes are still largely unknown. Here we describe Tn6188, a novel transposon in L. monocytogenes conferring tolerance to BC. Tn6188 is related to Tn554 from Staphylococcus (S. aureus and other Tn554-like transposons such as Tn558, Tn559 and Tn5406 found in various Firmicutes. Tn6188 comprises 5117 bp, is integrated chromosomally within the radC gene and consists of three transposase genes (tnpABC as well as genes encoding a putative transcriptional regulator and QacH, a small multidrug resistance protein family (SMR transporter putatively associated with export of BC that shows high amino acid identity to Smr/QacC from S. aureus and to EmrE from Escherichia coli. We screened 91 L. monocytogenes strains for the presence of Tn6188 by PCR and found Tn6188 in 10 of the analyzed strains. These isolates were from food and food processing environments and predominantly from serovar 1/2a. L. monocytogenes strains harboring Tn6188 had significantly higher BC minimum inhibitory concentrations (MICs (28.5 ± 4.7 mg/l than strains without Tn6188 (14 ± 3.2 mg/l. Using quantitative reverse transcriptase PCR we could show a significant increase in qacH expression in the presence of BC. QacH deletion mutants were generated in two L. monocytogenes strains and growth analysis revealed that ΔqacH strains had lower BC MICs than wildtype strains. In conclusion, our results provide evidence that Tn6188 is responsible for BC tolerance in various L

  7. 76 FR 63281 - Foreign-Trade Zone 78-Nashville, TN, Application for Subzone, Hemlock Semiconductor, L.L.C...

    Science.gov (United States)

    2011-10-12

    ... DEPARTMENT OF COMMERCE Foreign-Trade Zones Board [Docket 62-2011] Foreign-Trade Zone 78--Nashville, TN, Application for Subzone, Hemlock Semiconductor, L.L.C. (Polysilicon); Clarksville, TN An... polysilicon manufacturing facility of [[Page 63282

  8. Antibodies against the mono-methylated arginine-glycine repeat (MMA-RG) of the Epstein-Barr virus nuclear antigen 2 (EBNA2) identify potential cellular proteins targeted in viral transformation.

    Science.gov (United States)

    Ayoubian, Hiresh; Fröhlich, Thomas; Pogodski, Dagmar; Flatley, Andrew; Kremmer, Elisabeth; Schepers, Aloys; Feederle, Regina; Arnold, Georg J; Grässer, Friedrich A

    2017-08-01

    The Epstein-Barr virus is a human herpes virus with oncogenic potential. The virus-encoded nuclear antigen 2 (EBNA2) is a key mediator of viral tumorigenesis. EBNA2 features an arginine-glycine (RG) repeat at amino acids (aa)339-354 that is essential for the transformation of lymphocytes and contains symmetrically (SDMA) and asymmetrically (ADMA) di-methylated arginine residues. The SDMA-modified EBNA2 binds the survival motor neuron protein (SMN), thus mimicking SMD3, a cellular SDMA-containing protein that interacts with SMN. Accordingly, a monoclonal antibody (mAb) specific for the SDMA-modified RG repeat of EBNA2 also binds to SMD3. With the novel mAb 19D4 we now show that EBNA2 contains mono-methylated arginine (MMA) residues within the RG repeat. Using 19D4, we immune-precipitated and analysed by mass spectrometry cellular proteins in EBV-transformed B-cells that feature MMA motifs that are similar to the one in EBNA2. Among the cellular proteins identified, we confirmed by immunoprecipitation and/or Western blot analyses Aly/REF, Coilin, DDX5, FXR1, HNRNPK, LSM4, MRE11, NRIP, nucleolin, PRPF8, RBM26, SMD1 (SNRDP1) and THRAP3 proteins that are either known to contain MMA residues or feature RG repeat sequences that probably serve as methylation substrates. The identified proteins are involved in splicing, tumorigenesis, transcriptional activation, DNA stability and RNA processing or export. Furthermore, we found that several proteins involved in energy metabolism are associated with MMA-modified proteins. Interestingly, the viral EBNA1 protein that features methylated RG repeat motifs also reacted with the antibodies. Our results indicate that the region between aa 34-52 of EBNA1 contains ADMA or SDMA residues, while the region between aa 328-377 mainly contains MMA residues.

  9. Induction of the Tn10 Precise Excision in E. coli Cells after Accelerated Heavy Ions Irradiation

    CERN Document Server

    Zhuravel, D V

    2003-01-01

    The influence of the irradiation of different kinds on the indication of the structural mutations in the bacteria Escherichia coli is considered. The regularities of the Tn10 precise excision after accelerated ^{4}He and ^{12}C ions irradiations with different linear energy transfer (LET) were investigated. Dose dependences of the survival and relative frequency of the Tn10 precise excision were obtained. It was shown, that the relative frequency of the Tn10 precise excision is the exponential function from the irradiation dose. Relative biological efficiency (RBE), and relative genetic efficiency (RGE) were calculated, and were treated as the function of the LET.

  10. Monoclonal antibodies for treating cancer

    International Nuclear Information System (INIS)

    Dillman, R.O.

    1989-01-01

    The purpose of this study is to assess the current status of in-vivo use of monoclonal antibodies for treating cancer. Publications appearing between 1980 and 1988 were identified by computer searches using MEDLINE and CANCERLIT, by reviewing the table of contents of recently published journals, and by searching bibliographies of identified books and articles. More than 700 articles, including peer-reviewed articles and book chapters, were identified and selected for analysis. The literature was reviewed and 235 articles were selected as relevant and representative of the current issues and future applications for in-vivo monoclonal antibodies for cancer therapy and of the toxicity and efficacy which has been associated with clinical trials. Approaches include using antibody alone (interacting with complement or effector cells or binding directly with certain cell receptors) and immunoconjugates (antibody coupled to radioisotopes, drugs, toxins, or other biologicals). Most experience has been with murine antibodies. Trials of antibody alone and radiolabeled antibodies have confirmed the feasibility of this approach and the in-vivo trafficking of antibodies to tumor cells. However, tumor cell heterogeneity, lack of cytotoxicity, and the development of human antimouse antibodies have limited clinical efficacy. Although the immunoconjugates are very promising, heterogeneity and the antimouse immune response have hampered this approach as has the additional challenge of chemically or genetically coupling antibody to cytotoxic agents. As a therapeutic modality, monoclonal antibodies are still promising but their general use will be delayed for several years. New approaches using human antibodies and reducing the human antiglobulin response should facilitate treatment. 235 references

  11. A Phase I Study of Unimolecular Pentavalent (Globo-H-GM2-sTn-TF-Tn Immunization of Patients with Epithelial Ovarian, Fallopian Tube, or Peritoneal Cancer in First Remission

    Directory of Open Access Journals (Sweden)

    Roisin E. O’Cearbhaill

    2016-04-01

    Full Text Available We conducted a phase I study in ovarian cancer patients to evaluate the safety and immunogenicity of a synthetic unimolecular pentavalent carbohydrate vaccine (Globo-H, GM2, sTn, TF, and Tn supported on a peptide backbone, conjugated to keyhole limpet haemocyanin (KLH, and mixed with immunological adjuvant QS-21. Twenty-four advanced-stage, poor-risk, first-remission ovarian cancer patients were enrolled from January 2011–Septermber 2013. Three dose levels were planned (25, 50, 100 mcg with three cohorts of six patients each, with an additional 6-patient expansion cohort at the MTD. ELISA serologic IgM and IgG responses for each antigen was defined as positive response if antibody titers were ≥1:80 over the respective patient’s pre-vaccination serum. The study would be considered positive if at least four of 12 patients treated at the MTD showed immune responses for at least three of the five antigens. Twenty-four patients (median age, 54 years [range, 36–68] were included in the safety analysis. Histology was high-grade serous in 22 patients (92%; 18 had stage III and six stage IV disease. The vaccine was well-tolerated at all doses, with no DLTs. At the highest treated dose, IgG and/or IgM responses were recorded against ≥3 antigens in 9/12 patients (75%, ≥4 in 7/12 (58%, and 5 in 3/12 (25%. With a median follow-up of 19 months (range, 2–39, 20 patients (83% recurred and six (25% died. The unimolecular pentavalent vaccine construct was shown to be safe and immunogenic. Such a construct greatly simplifies regulatory requirements and manufacturing, facilitates scalability, and provides adaptability.

  12. EnviroAtlas - Memphis, TN - Meter-Scale Urban Land Cover (MULC) Data (2012)

    Data.gov (United States)

    U.S. Environmental Protection Agency — The Memphis, TN EnviroAtlas One Meter-scale Urban Land Cover (MULC) dataset comprises 2,733 km2 around the city of Memphis, surrounding towns, and rural areas. These...

  13. Internal size variations in Tn1546-like elements due to the presence of IS1216V

    DEFF Research Database (Denmark)

    Jensen, Lars Bogø

    1998-01-01

    to the presence of the IS sequence, IS1216 V. This IS sequence has previously been found integrated in Tn1546, Integration of the IS1216V element created both deletions and a duplication in a non-essential region of Tn1546. In several isolates, the entire vanY gene was deleted, proving that this gene is non......-essential for vancomycin resistance. (C) 1998 Federation of European Microbiological Societies....

  14. Characterization of Tn904 insertions in octopine Ti plasmid mutants of Agrobacterium tumefaciens.

    Science.gov (United States)

    Ooms, G; Klapwijk, P M; Poulis, J A; Schilperoort, R A

    1980-01-01

    Seven Tn904 insertion mutants of pTi Ach5 affecting Agrobacterium tumefaciens virulence were studied. The mutant character was shown to be plasmid borne. Four of these mutants were avirulent and carried an insertion in restriction endonuclease HpaI fragment 12, a 3.3-megadalton fragment, which therefore appears to be a Ti plasmid region essential for virulence. Two mutants were attenuated in virulence. The inserts mapped close to HpaI fragment 12. One mutant giving rise to small tumors with excessive adventitious root formation on Kalanchoe daigremontiana carried an insertion in the right side of the common sequence in the deoxyribonucleic acid of the Ti plasmid detected in crown gall tumors. The insertion behavior of Tn904 was studied by analyzing 11 independently isolated and randomly chosen mutants. The Tn904 inserts did not affect oncogenicity, tumor morphology, bacterial transfer functions, octopine catabolism functions, or vital parts of the Ti plasmid, such as the origin of replication. Most of the Tn904 inserts were concentrated in a small part of the map. The size of additional deoxyribonucleic acid as a result of Tn904 inserts varied between 5 and 15 megadaltons. In two cases a Ti plasmid was found with two Tn904 insertions at different positions. Images PMID:6252198

  15. Antibody biotechnology

    African Journals Online (AJOL)

    STORAGESEVER

    2009-07-06

    Jul 6, 2009 ... and automated, the hybrid cells can be stored for many years in liquid nitrogen and antibodies production is homogeneous. The hybridoma method .... they may be modified to vehicle active molecules such as radio-isotopes, toxins, cytokines, enzyme etc. In these cases, the therapeutic effect is due to ...

  16. Catalytic Antibodies

    Indian Academy of Sciences (India)

    The ability of the highly evolved machinery of immune system to produce structurally and functionally complex ... to Pauling, if the structure of the antigen binding site of antibodies were to be produced in a random ..... where the immune system of the body is destructive, as in autoimmune disorders or after organ transplant.

  17. Catalytic Antibodies

    Indian Academy of Sciences (India)

    While chemistry provides the framework for understanding the structure and function of biomolecules, the immune sys- tem provides a highly evolved natural process to generate one class of complex biomolecules – the antibodies. A combination of the two could be exploited to generate new classes of molecules with novel ...

  18. Variation on a theme; an overview of the Tn916 / Tn1545 family of mobile genetic elements in the oral and nasopharyngeal streptococci.

    Directory of Open Access Journals (Sweden)

    Francesco eSantoro

    2014-10-01

    Full Text Available The oral and nasopharyngeal streptococci are a major part of the normal microbiota in humans. Most human associated streptococci are considered commensals however a small number of them are pathogenic, causing a wide range of diseases including oral infections such as dental caries and periodontitis and diseases at other body sites including sinusitis and endocarditis, and in the case of Streptococcus pneumoniae, meningitis. Both phenotypic and sequence based studies have shown that the human associated streptococci from the mouth and nasopharynx harbour a large number of antibiotic resistance genes and these are often located on mobile genetic elements known as conjugative transposons or integrative and conjugative elements of the Tn916 / Tn1545 family. These mobile genetic elements are responsible for the spread of the resistance genes between streptococci and also between streptococci and other bacteria. In this review we describe the resistances conferred by, and the genetic variations between the many different Tn916-like elements found in recent studies of oral and nasopharyngeal streptococci and show that Tn916-like elements are important mediators of antibiotic resistance genes within this genus. We will also discuss the role of the oral environment and how this is conducive to the transfer of these elements and discuss the contribution of both transformation and conjugation on the transfer and evolution of these elements in different streptococci.

  19. Identification of a novel linear B-cell epitope using a monoclonal antibody against the carboxy terminus of the canine distemper virus nucleoprotein and sequence analysis of the identified epitope in different CDV isolates.

    Science.gov (United States)

    Yi, Li; Cao, Zhigang; Tong, Mingwei; Cheng, Yuening; Yang, Yong; Li, Shuang; Wang, Jianke; Lin, Peng; Sun, Yaru; Zhang, Miao; Cheng, Shipeng

    2017-09-29

    The Nucleoprotein (NP) is the most abundant and highly immunogenic protein in canine distemper virus (CDV), playing an important role in CDV viral replication and assembly. In this study, a specific monoclonal antibody, named C8, was produced against the NP protein C terminal (amino acids 401-523). A linear N protein epitope was identified by subjecting a series of partially overlapping synthesized peptides to enzyme-linked immunosorbent assay (ELISA) analysis.The results indicated that 444 GDKYPIHFNDER 455 was the minimal linear epitope that could be recognized by mAb C8. Sequence alignments demonstrated that this linear epitope is less conserved among three CDV genotypes. We next analyzed the level of conservation of the defined epitope in19 Chinese CDV clinical isolates, and it has one site variation in amino acid among these CDV isolations. 2 isolates have the amino acid mutations F451L, while one has P448Ssubstitution.Phylogenetic analysis showed the two isolates with F451Lsubstitution had a closer relationship in a virulent strain ZJ-7, so the epitope may be a significant tag associated with virus virulence. This collection of mAb along with defined linear epitope may provide useful reagents for investigations of NP protein function and the development of CDV specific diagnostics.

  20. Peri-operative troponin monitoring using a prototype high-sensitivity cardiac troponin I (hs-cTnI) assay: comparisons with hs-cTnT and contemporary cTnI assays.

    LENUS (Irish Health Repository)

    Lee, Graham R

    2013-09-18

    Non-cardiac surgery is associated with major vascular complications and higher incidences of elevated plasma troponin (cTn) concentration. Goal-directed therapy (GDT) is a stroke volume (SV)-guided approach to intravenous (IV) fluid therapy that improves tissue perfusion, oxygenation and reduces post-operative complications. In patients undergoing major gastro-intestinal surgery, we compared high sensitive and contemporary troponin assays and correlated results with patient outcome.

  1. Myofilament Calcium Sensitivity: Mechanistic Insight into TnI Ser-23/24 and Ser-150 Phosphorylation Integration

    Directory of Open Access Journals (Sweden)

    Hussam E Salhi

    2016-12-01

    Full Text Available Troponin I (TnI is a major regulator of cardiac muscle contraction and relaxation. During physiological and pathological stress, TnI is differentially phosphorylated at multiple residues through different signaling pathways to match cardiac function to demand. The combination of these TnI phosphorylations can exhibit an expected or unexpected functional integration, whereby the function of two phosphorylations are different than that predicted from the combined function of each individual phosphorylation alone. We have shown that TnI Ser-23/24 and Ser-150 phosphorylation exhibit functional integration and are simultaneously increased in response to cardiac stress. In the current study, we investigated the functional integration of TnI Ser-23/24 and Ser-150 to alter cardiac contraction. We hypothesized that Ser-23/24 and Ser-150 phosphorylation each utilize distinct molecular mechanisms to alter the TnI binding affinity within the thin filament. Mathematical modeling predicts that Ser-23/24 and Ser-150 phosphorylation affect different TnI affinities within the thin filament to distinctly alter the Ca2+-binding properties of troponin. Protein binding experiments validate this assertion by demonstrating pseudo-phosphorylated Ser-150 decreases the affinity of isolated TnI for actin, whereas Ser-23/24 pseudo-phosphorylation is not different from unphosphorylated. Thus, our data supports that TnI Ser-23/24 affects TnI-TnC binding, while Ser-150 phosphorylation alters TnI-actin binding. By measuring force development in troponin-exchanged skinned myocytes, we demonstrate that the Ca2+ sensitivity of force is directly related to the amount of phosphate present on TnI. Furthermore, we demonstrate that Ser-150 pseudo-phosphorylation blunts Ser-23/24-mediated decreased Ca2+-sensitive force development whether on the same or different TnI molecule. Therefore, TnI phosphorylations can integrate across troponins along the myofilament. These data demonstrate

  2. Nucleotide sequence and functional analysis of the tet (M)-carrying conjugative transposon Tn5251 of Streptococcus pneumoniae.

    Science.gov (United States)

    Santoro, Francesco; Oggioni, Marco R; Pozzi, Gianni; Iannelli, Francesco

    2010-07-01

    The Tn916-like genetic element Tn5251 is part of the composite conjugative transposon (CTn) Tn5253 of Streptococcus pneumoniae, a 64.5-kb chromosomal element originally called Omega(cat-tet) BM6001. DNA sequence analysis showed that Tn5251 is 18 033-bp long and contains 22 ORFs, 20 of which have the same direction of transcription. Annotation was possible for 11 out of 22 ORFs, including the tet(M) tetracycline resistance gene and int and xis involved in the integration/excision process. Autonomous copies of Tn5251 were generated during matings of Tn5253-containing donors with S. pneumoniae and Enterococcus faecalis. Tn5251 was shown to integrate at different sites in the bacterial chromosome. It behaves as a fully functional CTn capable of independent conjugal transfer to a variety of bacterial species including S. pneumoniae, Streptococcus gordonii, Streptococcus pyogenes, Streptococcus agalactiae, E. faecalis and Bacillus subtilis. The excision of Tn5251 produces a circular intermediate and a deletion in Tn5253 at a level of 1.2 copies per 10(5) chromosomes.

  3. Novel Tn4371-ICE like element in Ralstonia pickettii and genome mining for comparative elements.

    Science.gov (United States)

    Ryan, Michael P; Pembroke, J Tony; Adley, Catherine C

    2009-11-26

    Integrative Conjugative Elements (ICEs) are important factors in the plasticity of microbial genomes. An element related to the ICE Tn4371 was discovered during a bioinformatic search of the Ralstonia pickettii 12J genome. This element was analysed and further searches carried out for additional elements.A PCR method was designed to detect and characterise new elements of this type based on this scaffold and a culture collection of fifty-eight Ralstonia pickettii and Ralstonia insidiosa strains were analysed for the presence of the element. Comparative sequence analysis of bacterial genomes has revealed the presence of a number of uncharacterised Tn4371-like ICEs in the genomes of several beta and gamma- Proteobacteria. These elements vary in size, GC content, putative function and have a mosaic-like structure of plasmid- and phage-like sequences which is typical of Tn4371-like ICEs. These elements were found after a through search of the GenBank database. The elements, which are found in Ralstonia, Delftia, Acidovorax, Bordetella, Comamonas, Acidovorax, Congregibacter, Shewanella, Pseudomonas Stenotrophomonas, Thioalkalivibrio sp. HL-EbGR7, Polaromonas, Burkholderia and Diaphorobacter sp. share a common scaffold. A PCR method was designed (based on the Tn4371- like element detected in the Ralstonia pickettii 12J genome) to detect and characterise new elements of this type. All elements found in this study possess a common scaffold of core genes but contain different accessory genes. A new uniform nomenclature is suggested for ICEs of the Tn4371 family. Two novel Tn4371-like ICE were discovered and characterised, using the novel PCR method described in two different isolates of Ralstonia pickettii from laboratory purified water.

  4. The VanE operon in Enterococcus faecalis N00-410 is found on a putative integrative and conjugative element, Tn6202.

    Science.gov (United States)

    Boyd, D A; Mulvey, M R

    2013-02-01

    To date no complete genetic structure of acquired DNA harbouring a d-Ala-d-Ser operon in an Enterococcus is known. We wished to characterize the acquired DNA harbouring the vanE operon located in the Enterococcus faecalis N00-410 chromosome. Whole genome sequencing of E. faecalis N00-410 was conducted by massively parallel sequencing. Two sequence contigs harbouring the vanE region were linked by PCR and the acquired DNA harbouring the vanE operon was completely characterized. Excision/integration of the region was determined by PCR and transfer attempted by conjugation. The regions flanking the vanE operon were analysed and a total of 42 open reading frames were identified in a region flanked by inverted terminal and direct repeats (Tn6202). Tn6202 could be excised from the chromosome, circularized and the target site rejoined, but transfer could not be demonstrated. The vanE operon was found on the putative integrative and conjugative element Tn6202 in the E. faecalis N00-410 chromosome. This represents the first characterization of acquired DNA harbouring a D-Ala-D-Ser operon.

  5. Production of diketopiperazine derivative cyclo (l-leu-l-arg) by streptomyces sp. tn262 after exposure to heat-killed fungus fusarium sp

    International Nuclear Information System (INIS)

    Elleuch, L.; Smaoui, S.; Najah, S.; Sellem, I

    2013-01-01

    In a screening program for new active secondary metabolites producers, a strain of Streptomyces called TN262 was isolated from Tunisian soil and selected for its ability to produce eleven active compounds in pure culture conditions. In this work, the effect of different concentrations of heat-killed fungus Fusarium sp. on the production of active compounds by TN262 strain was studied. The ethyl acetate extract from the culture of Streptomyces sp. TN262 combined with heat-killed Fusarium sp. at 50 micro g/ml inhibited the growth of the three used indicator microorganisms. In fact, an increase of 36%, 21% and 20% in inhibitory activity was obtained against Micrococcus luteus LB 14110, Escherichia coli ATCC 8739 and Fusarium sp. respectively. The HPLC chromatographic profiles of the ethyl acetate extracts from both culture conditions were different and an additional active compound was produced only under induced conditions. This active component was isolated and identified as Cyclo (L-Leu-L-Arg) (1), a diketopiperazine derivative, possessing antibacterial and antifungal activity. Consequently, this study showed that the addition of heat-killed fungus is a useful method for inducing the production of bioactive compounds. (author)

  6. Introduction of transposon Tn901 into a plasmid of Anacystis nidulans: preparation for cloning in cyanobacteria

    NARCIS (Netherlands)

    van den Hondel, C. A.; Verbeek, S.; van der Ende, A.; Weisbeek, P. J.; Borrias, W. E.; van Arkel, G. A.

    1980-01-01

    We have used the TEM beta-lactamase transposon Tn901, located on Escherichia coli plasmid pRI46, to introduce in vivo a genetic marker into plasmid pUH24, present in the cyanobacterial strain Anacystis nidulans R-2. Restriction enzyme analysis and heteroduplex studies of the 8.3 x 10(6)-dalton

  7. Characterization of glucansucrase and dextran from Weissella sp. TN610 with potential as safe food additives.

    Science.gov (United States)

    Bejar, Wacim; Gabriel, Valérie; Amari, Myriam; Morel, Sandrine; Mezghani, Monia; Maguin, Emmanuelle; Fontagné-Faucher, Catherine; Bejar, Samir; Chouayekh, Hichem

    2013-01-01

    Pear-derived Weissella sp. TN610 produced extracellular glycosyltransferase activity responsible for the synthesis of soluble exopolysaccharide from sucrose. Acid and dextranase-catalyzed hydrolysis revealed that the synthesized polymer was a glucan. According to (1)H and (13)C NMR analysis, the glucan produced by TN610 was a linear dextran made of 96% α-(1→6) and 4% α-(1→3) linkages. Zymogram analysis confirmed the presence of a unique glucansucrase of approximately 180 kDa in the cell-free supernatant from TN610. The crude enzyme, optimally active at 37°C and pH 5, has promising potential for application as a food additive since it catalyzes dextran synthesis in sucrose-supplemented milk, allowing its solidification. A 4257-bp product corresponding to the mature glucansucrase gene was amplified by PCR from TN610. It encoded a polypeptide of 1418 residues having a calculated molecular mass of 156.089 kDa and exhibiting 96% and 95% identity with glucansucrases from Lactobacillus fermentum Kg3 and Weissella cibaria CMU, respectively. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Mini-Tn7 transposons for site-specific tagging of bacteria with fluorescent proteins

    DEFF Research Database (Denmark)

    Lambertsen, L.; Sternberg, Claus; Molin, Søren

    2004-01-01

    the Escherichia coli lac derived promoter, P-A1/04/03, or from the growth-rate-dependent Escherichia coli promoter P-rrnB P1. The mini-Tn7 transposons were inserted and tested in the soil bacterium, Pseudomonas putida KT2440. Successful and site-specific tagging was verified by Southern blots as well as by PCR...

  9. 77 FR 17360 - Proposed Amendment of Class E Airspace; Memphis, TN

    Science.gov (United States)

    2012-03-26

    ... regulations to assign the use of airspace necessary to ensure the safety of aircraft and the efficient use of... Branch, MS, Olive Branch Airport (Lat. 34[deg]58'44'' N., long. 89[deg]47'13'' W.) General DeWitt Spain... radius of General DeWitt Spain Airport; excluding that airspace within the Millington, TN, Class E...

  10. Morphological and functional diversity of ectomycorrhizal fungi on Roan Mountain (NC/TN)

    Science.gov (United States)

    Claire Bird; Coleman McCleneghan

    2005-01-01

    A comparison of ectomycorrhizal morphotypes and hypogeous fungi (truffles and false-truffles) in northern hardwood and spruce-fir forests on Roan Mountain (NC/TN) was performed to increase our knowledge of the fungal communities in the Southern Appalachian high elevation forests. These forests are home to an endangered subspecies and mycophagist, the Carolina northern...

  11. 75 FR 69398 - Foreign-Trade Zone 78-Nashville, TN; Application for Expansion

    Science.gov (United States)

    2010-11-12

    ... DEPARTMENT OF COMMERCE Foreign-Trade Zones Board [Docket 64-2010] Foreign-Trade Zone 78--Nashville, TN; Application for Expansion An application has been submitted to the Foreign-Trade Zones Board (the... application was submitted pursuant to the provisions of the Foreign- Trade Zones Act, as amended (19 U.S.C...

  12. 76 FR 39379 - Expansion of Foreign-Trade Zone 78; Nashville, TN

    Science.gov (United States)

    2011-07-06

    ... DEPARTMENT OF COMMERCE Foreign-Trade Zones Board [Order No. 1768] Expansion of Foreign-Trade Zone 78; Nashville, TN Pursuant to its authority under the Foreign-Trade Zones Act of June 18, 1934, as amended (19 U.S.C. 81a-81u), the Foreign-Trade Zones Board (the Board) adopts the following Order: Whereas...

  13. EFFECT TN EWES OF OESTROGEN PRIMING AND GnRH ON LH ...

    African Journals Online (AJOL)

    EFFECT TN EWES OF OESTROGEN PRIMING AND GnRH ON LH RELEASE. AND LUTEAL FUNCTION DURING EARLY LACTATION IN SPRING. Receipt of MS 22-03-1979. C.D. Hamilton*, A.W. Lishman and P.A. Lamb. Department ol Animal Science, Universitlt ol'Natol, nercrmaitzburg, 3200. (Key words. Oestrogen ...

  14. Molecular analysis of Tn1546 in Enterococcus faecium isolated from animals and humans

    DEFF Research Database (Denmark)

    Jensen, Lars Bogø; Ahrens, Peter; Dons, L.

    1998-01-01

    The internal areas and the position of integration of the glycopeptide resistance element Tn1546 were characterized by using PCR fragment length polymorphism, sequencing, and DNA hybridization techniques with 38 high-level vancomycin-resistant Enterococcus faecium isolates of human and animal...

  15. 76 FR 35909 - Temporary Concession Contract for Big South Fork National Recreation Area, TN/KY

    Science.gov (United States)

    2011-06-20

    ... years. The visitor services include providing backcountry lodging accommodations, food and beverage, and... DEPARTMENT OF THE INTERIOR National Park Service [[NPS-WASO-CONC-0511-7144; 2410-OYC] Temporary Concession Contract for Big South Fork National Recreation Area, TN/KY AGENCY: National Park Service...

  16. Induction of transposon TN1 translocation under the action of different mutagens

    International Nuclear Information System (INIS)

    Kubanejshvili, M.G.; Smirnov, S.P.; Tarasov, V.A.

    1983-01-01

    Migration of ampicillin transposon Tn1 under normal conditions in Escherichia coli cells proceeds with low frequency (10 -4 transpositions for cell). The low transposition frequency is conditioned by the transposition repression, realized by the gene-repressor in transposon structure and, probably, by other regulating genes of the bacterium-host. E. coli cell treatment by physical and chemical mutagens resulted in induction of translocation of ampicillin transposon Tn1 from plasmid RP4 into other replicons. Mitomycin C and ultraviolet radiation produced stronger inducing effect as compared to nitroso-guanidine (NG). The effect of the given mutagens on transposition Tn1 correlated with their activating capacity with respect to inducible SOS-functions of E coli. The mutation of rec A didn't influence on spontaneous Tn1 transposition, but blocked completely the induction of transposition process under mutagen effect. The relationship of inducible transposition with SOS-functions in E. coli cells, controlled by recA and lexA genes, as well as the possible role of the process in genetic microorganism variability are discussed in the paper

  17. 75 FR 52364 - Notice of Intent To Repatriate Cultural Items: Memphis Pink Palace Museum, Memphis, TN

    Science.gov (United States)

    2010-08-25

    ...: Memphis Pink Palace Museum, Memphis, TN AGENCY: National Park Service, Interior. ACTION: Notice. Notice is....S.C. 3005, of the intent to repatriate cultural items in the possession of the Memphis Pink Palace... site. Officials of the Memphis Pink Palace Museum have determined that, pursuant to 25 U.S.C. 3001(3)(B...

  18. 75 FR 61696 - Foreign-Trade Zone 148-Knoxville, TN; Application for Subzone; Toho Tenax America, Inc. (Carbon...

    Science.gov (United States)

    2010-10-06

    ...--Knoxville, TN; Application for Subzone; Toho Tenax America, Inc. (Carbon Fiber and Oxidized Polyacrylonitrile Fiber Manufacturing); Rockwood, TN An application has been submitted to the Foreign-Trade Zones..., requesting special-purpose subzone status for the carbon fiber and oxidized polyacrylonitrile fiber (OPF...

  19. 77 FR 13073 - Designation for the Jamestown, ND; Lincoln, NE; Memphis, TN; and Sioux City, IA Areas

    Science.gov (United States)

    2012-03-05

    ... the Jamestown, ND; Lincoln, NE; Memphis, TN; and Sioux City, IA Areas AGENCY: Grain Inspection..., IA areas, Lincoln, Midsouth, and Sioux City, respectively were the sole applicants for designation to.../2015 Midsouth Memphis, TN (901) 942-3216 4/1/2012 3/31/2015 Sioux City Sioux City, IA......... (712...

  20. Large-Scale Low-Cost NGS Library Preparation Using a Robust Tn5 Purification and Tagmentation Protocol

    Directory of Open Access Journals (Sweden)

    Bianca P. Hennig

    2018-01-01

    Full Text Available Efficient preparation of high-quality sequencing libraries that well represent the biological sample is a key step for using next-generation sequencing in research. Tn5 enables fast, robust, and highly efficient processing of limited input material while scaling to the parallel processing of hundreds of samples. Here, we present a robust Tn5 transposase purification strategy based on an N-terminal His6-Sumo3 tag. We demonstrate that libraries prepared with our in-house Tn5 are of the same quality as those processed with a commercially available kit (Nextera XT, while they dramatically reduce the cost of large-scale experiments. We introduce improved purification strategies for two versions of the Tn5 enzyme. The first version carries the previously reported point mutations E54K and L372P, and stably produces libraries of constant fragment size distribution, even if the Tn5-to-input molecule ratio varies. The second Tn5 construct carries an additional point mutation (R27S in the DNA-binding domain. This construct allows for adjustment of the fragment size distribution based on enzyme concentration during tagmentation, a feature that opens new opportunities for use of Tn5 in customized experimental designs. We demonstrate the versatility of our Tn5 enzymes in different experimental settings, including a novel single-cell polyadenylation site mapping protocol as well as ultralow input DNA sequencing.

  1. Identification of Pathogenicity-Related Genes in Biofilm-Defective Acidovorax citrulli by Transposon Tn5 Mutagenesis

    Directory of Open Access Journals (Sweden)

    Jinyan Luo

    2015-11-01

    Full Text Available Biofilm formation is important for virulence of a large number of plant pathogenic bacteria. Indeed, some virulence genes have been found to be involved in the formation of biofilm in bacterial fruit blotch pathogen Acidovorax citrulli. However, some virulent strains of A. citrulli were unable to format biofilm, indicating the complexity between biofilm formation and virulence. In this study, virulence-related genes were identified in the biofilm-defective strain A1 of A. citrulli by using Tn5 insertion, pathogenicity test, and high-efficiency thermal asymmetric interlaced PCR (hiTAIL-PCR. Results from this study indicated that 22 out of the obtained 301 mutants significantly decreased the virulence of strain A1 compared to the wild-type. Furthermore, sequence analysis indicated that the obtained 22 mutants were due to the insertion of Tn5 into eight genes, including Aave 4244 (cation diffusion facilitator family transporter, Aave 4286 (hypothetical protein, Aave 4189 (alpha/beta hydrolase fold, Aave 1911 (IMP dehydrogenase/GMP reductase domain, Aave 4383 (bacterial export proteins, family 1, Aave 4256 (Hsp70 protein, Aave 0003 (histidine kinase, DNA gyrase B, and HSP90-like ATPase, and Aave 2428 (pyridoxal-phosphate dependent enzyme. Furthermore, the growth of mutant Aave 2428 was unaffected and even increased by the change in incubation temperature, NaCl concentration and the pH of the LB broth, indicating that this gene may be directly involved in the bacterial virulence. Overall, the determination of the eight pathogenicity-related genes in strain A1 will be helpful to elucidate the pathogenesis of biofilm-defective A. citrulli.

  2. Replacing reprogramming factors with antibodies selected from combinatorial antibody libraries.

    Science.gov (United States)

    Blanchard, Joel W; Xie, Jia; El-Mecharrafie, Nadja; Gross, Simon; Lee, Sohyon; Lerner, Richard A; Baldwin, Kristin K

    2017-10-01

    The reprogramming of differentiated cells into induced pluripotent stem cells (iPSCs) is usually achieved by exogenous induction of transcription by factors acting in the nucleus. In contrast, during development, signaling pathways initiated at the membrane induce differentiation. The central idea of this study is to identify antibodies that can catalyze cellular de-differentiation and nuclear reprogramming by acting at the cell surface. We screen a lentiviral library encoding ∼100 million secreted and membrane-bound single-chain antibodies and identify antibodies that can replace either Sox2 and Myc (c-Myc) or Oct4 during reprogramming of mouse embryonic fibroblasts into iPSCs. We show that one Sox2-replacing antibody antagonizes the membrane-associated protein Basp1, thereby de-repressing nuclear factors WT1, Esrrb and Lin28a (Lin28) independent of Sox2. By manipulating this pathway, we identify three methods to generate iPSCs. Our results establish unbiased selection from autocrine combinatorial antibody libraries as a robust method to discover new biologics and uncover membrane-to-nucleus signaling pathways that regulate pluripotency and cell fate.

  3. Tn5/7-lux: a versatile tool for the identification and capture of promoters in gram-negative bacteria.

    Science.gov (United States)

    Bruckbauer, Steven T; Kvitko, Brian H; Karkhoff-Schweizer, RoxAnn R; Schweizer, Herbert P

    2015-02-04

    The combination of imaging technologies and luciferase-based bioluminescent bacterial reporter strains provide a sensitive and simple non-invasive detection method (photonic bioimaging) for the study of diverse biological processes, as well as efficacy of therapeutic interventions, in live animal models of disease. The engineering of bioluminescent bacteria required for photonic bioimaging is frequently hampered by lack of promoters suitable for strong, yet stable luciferase gene expression. We devised a novel method for identification of constitutive native promoters in Gram-negative bacteria. The method is based on a Tn5/7 transposon that exploits the unique features of Tn5 (random transposition) and Tn7 (site-specific transposition). The transposons are designed such that Tn5 transposition will allow insertion of a promoter-less bacterial luxCDABE operon downstream of a bacterial gene promoter. Cloning of DNA fragments from luminescent isolates results in a plasmid that replicates in pir (+) hosts. Sequencing of the lux-chromosomal DNA junctions on the plasmid reveals transposon insertion sites within genes or operons. The plasmid is also a mini-Tn7-lux delivery vector that can be used to introduce the promoter-lux operon fusion into other derivatives of the bacterium of interest in an isogenic fashion. Alternatively, promoter-containing sequences can be PCR-amplified from plasmid or chromosomal DNA and cloned into a series of accompanying mini-Tn7-lux vectors. The mini-Tn5/7-lux and mini-Tn7-lux vectors are equipped with diverse selection markers and thus applicable in numerous Gram-negative bacteria. Various mini-Tn5/7-lux vectors were successfully tested for transposition and promoter identification by imaging in Acinetobacter baumannii, Escherichia coli, and Burkholderia pseudomallei. Strong promoters were captured for lux expression in E. coli and A. baumannii. Some mini-Tn7-lux vectors are also equipped with attB sites for swapping of the lux operon with

  4. Effect of anti-carbohydrate antibodies on HIV infection in a monocytic cell line (U937)

    DEFF Research Database (Denmark)

    Hansen, J E; Nielsen, C; Clausen, H

    1991-01-01

    Monoclonal antibodies (mAbs) against carbohydrate epitopes of gp120 have recently been found to inhibit HIV infection of lymphocytes in vitro thereby opening new possibilities for vaccine considerations. Antibody-dependent enhancement of infection has however come increasingly into focus....... This study therefore investigated the neutralization of HIV in a monocytic cell line (U937) using mAbs against these carbohydrate gp120-epitopes. While antibodies against one of the epitopes (AI) neutralized infection of U937 cells despite binding to the Fc-receptor, one mAb against the sialosyl-Tn epitope...... enhanced infection. This enhancement was independent of complement and could be blocked by mAb Leu3a against the CD4-receptor. The study indicated that enhancement of infection in monocytic cells can occur by the same anti-carbohydrate antibodies that neutralize infection in lymphocytes, and that antibody...

  5. Identification of Novel Breast Cancer Antigens Using Phage Antibody Libraries

    National Research Council Canada - National Science Library

    Marks, James

    2002-01-01

    The purpose of this project is to use phage antibody libraries to identify novel breast tumor antigens The antibodies could be used for breast cancer immunotherapy and the antigens could be used as cancer vaccines...

  6. Diversity and Evolution of the Tn5801-tet(M)-Like Integrative and Conjugative Elements among Enterococcus, Streptococcus, and Staphylococcus.

    Science.gov (United States)

    León-Sampedro, Ricardo; Novais, Carla; Peixe, Luísa; Baquero, Fernando; Coque, Teresa M

    2016-01-04

    This work describes the diversity and evolution of Tn5801 among enterococci, staphylococci, and streptococci based on analysis of the 5,073 genomes of these bacterial groups available in gene databases. We also examined 610 isolates of Enterococcus (from 10 countries, 1987 to 2010) for the presence of this and other known CTn-tet(M) elements due to the scarcity of data about Tn5801 among enterococci. Genome location (by ICeu-I-pulsed-field gel electrophoresis [PFGE] hybridization/integration site identification), conjugation and fitness (by standard methods), Tn5801 characterization (by long-PCR mapping/sequencing), and clonality (by PFGE/multilocus sequence typing [MLST]) were studied. Twenty-three Tn5801 variants (17 unpublished) clustered in two groups, designated "A" (25 kb; n = 14; predominant in Staphylococcus aureus) and "B" (20 kb; n = 9; predominant in Streptococcus agalactiae). The percent GC content of the common backbone suggests a streptococcal origin of Tn5801 group B, with further acquisition of a 5-kb fragment that resulted in group A. Deep sequence analysis allowed identification of variants associated with clonal lineages of S. aureus (clonal complex 8 [CC8], sequence type 239 [ST239]), S. agalactiae (CC17), Enterococcus faecium (ST17/ST18), or Enterococcus faecalis (ST8), local variants, or variants located in different species and geographical areas. All Tn5801 elements were chromosomally located upstream of the guaA gene, which serves as an integration hot spot. Transferability was demonstrated only for Tn5801 type B among E. faecalis clonal backgrounds, which eventually harbored another Tn5801 copy. The study documents early acquisition of Tn5801 by Enterococcus, Staphylococcus, and Streptococcus. Clonal waves of these pathogens seem to have contributed to the geographical spread and local evolution of the transposon. Horizontal transfer, also demonstrated, could explain the variability observed, with the isolates often containing sequences of

  7. The TN-GEMINI: experience on a versatile alpha waste transport container

    International Nuclear Information System (INIS)

    Roland, V.; Chanzy, Y.

    2001-01-01

    The present paper discusses experience gained in moving alpha wastes and its teachings regarding transport aspects of D and D. Alpha wastes are generated in fuel cycle facilities such as those involved in reprocessing, in manufacture of mixed oxide fuel, and by research laboratories. If a significant amount of wastes has to be transported, then a Type B packaging is required. Developed by Transnucleaire and COGEMA, the TN GEMINI container enables nuclear facilities operators to optimise their alpha waste transport management, and more generally contribute to their D and D projects. After describing succinctly the design of the TN GEMINI, the paper will explain how the packaging is being operated. Teachings from experience will be shared. (orig.)

  8. [Effects of Total Nitrogen and BOD5/TN on Anaerobic Ammonium Oxidation-Denitrification Synergistic Interaction of Mature Landfill Leachate in Aged Refuse Bioreactor].

    Science.gov (United States)

    Yang, Ying-ying; Chen, Yi; Lj, Ming-jie; Xie, Bing

    2015-04-01

    Mature landfill leachate, featured with high ammonium (NH4+) content and low biodegrade ability (low BOD5/COD ratio), is hard to be treated. This study mainly focused on the effects of influent TN (total nitrogen) loading and BOD5/TN ratios on the nitrogen removal efficiency of landfill leachate by landfill bioreactors. The results showed that when the influent total nitrogen loading was in the range of 15 g x (m3 x d)(-1) to 25 g x (m3 x d)(-1), the TN removal loading could remain stable between 10 g x (m3 x d)(-1) and 12 g x (m3 x d)(-1), while the TN removal efficiency decreased from 67.7% to 60.2% with the increasing loading. Therefore, TN loading shocks would lower the bioreactor's TN removal rate, but would not affect its TN removal loading. When the influent BOD5/TN ratio was increased from 0.3 to 0.4 and the TN loading was controlled at 9 g x (m3 x d)(-1), the TN removal rates were increased from 79.9% to 89.9% and 86.2% in anaerobic and aerobic, respectively. This implied that properly enhancing BOD5/TN ratio could significantly increase the TN removal efficiency of the bioreactor, and the effect was more significant under anaerobic condition. Analysis of nitrogen removal pathways showed that denitrification and anammox could take place synergistically in landfill bioreactor.

  9. Design analysis report for the TN-WHC cask and transportation system

    Energy Technology Data Exchange (ETDEWEB)

    Brisbin, S.A., Fluor Daniel Hanford

    1997-02-13

    This document presents the evaluation of the Spent Nuclear Fuel Cask and Transportation System. The system design was developed by Transnuclear, Inc. and its team members NAC International, Nelson Manufacturing, Precision Components Corporation, and Numatec, Inc. The cask is designated the TN-WHC cask. This report describes the design features and presents preliminary analyses performed to size critical dimensions of the system while meeting the requirements of the performance specification.

  10. Monoclonal antibodies against rat leukocyte surface antigens

    NARCIS (Netherlands)

    van den Berg, T. K.; Puklavec, M. J.; Barclay, A. N.; Dijkstra, C. D.

    2001-01-01

    Monoclonal antibodies have proven to be powerful tools for studying the properties of leukocyte surface antigens and the cells that express them. In the past decades many monoclonal antibodies (mAb) for identifying the different rat leukocyte surface antigens have been described. A list of mAb is

  11. 77 FR 17408 - Foreign-Trade Zone 204-Tri-Cities Area, TN/VA; Application for Reorganization Under Alternative...

    Science.gov (United States)

    2012-03-26

    ... Technology Park, 4509 West Stone Drive, Kingsport, Hawkins County, TN; Site 9 (134 acres, expires 6/30/15.... The grantee's proposed service area under the ASF would be the Counties of Sullivan, Hawkins, Greene...

  12. Acetylcholine receptor antibody

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003576.htm Acetylcholine receptor antibody To use the sharing features on this page, please enable JavaScript. Acetylcholine receptor antibody is a protein found in the blood ...

  13. Platelet antibodies blood test

    Science.gov (United States)

    This blood test shows if you have antibodies against platelets in your blood. Platelets are a part of the blood ... Chernecky CC, Berger BJ. Platelet antibody - blood. In: Chernecky ... caused by platelet destruction, hypersplenism, or hemodilution. ...

  14. Opposites attract in bispecific antibody engineering

    NARCIS (Netherlands)

    van Gils, Marit J.; Sanders, Rogier W.

    2017-01-01

    Bispecific antibodies show great promise as intrinsic combination therapies, but often suffer from poor physiochemical properties, many times related to poor heterodimerization. De Nardis et al. identify specific electrostatic interactions that facilitate efficient heterodimerization, resulting in

  15. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    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.

  16. Antibody profiling sensitivity through increased reporter antibody layering

    International Nuclear Information System (INIS)

    Apel, William A.; Thompson, Vickie S.

    2013-01-01

    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.

  17. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    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.

  18. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    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.

  19. Antibody profiling sensitivity through increased reporter antibody layering

    Energy Technology Data Exchange (ETDEWEB)

    Apel, William A.; Thompson, Vicki S.

    2017-03-28

    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.

  20. Dissemination and Persistence of blaCTX-M-9 Are Linked to Class 1 Integrons Containing CR1 Associated with Defective Transposon Derivatives from Tn402 Located in Early Antibiotic Resistance Plasmids of IncHI2, IncP1-α, and IncFI Groups

    Science.gov (United States)

    Novais, Ângela; Cantón, Rafael; Valverde, Aránzazu; Machado, Elisabete; Galán, Juan-Carlos; Peixe, Luísa; Carattoli, Alessandra; Baquero, Fernando; Coque, Teresa M.

    2006-01-01

    This study analyzes the diversity of In60, a class 1 integron bearing CR1 and containing blaCTX-M-9, and its association with Tn402, Tn21, and classical conjugative plasmids among 45 CTX-M-9-producing clinical strains (41 Escherichia coli strains, 2 Klebsiella pneumoniae strains, 1 Salmonella enterica strain, and 1 Enterobacter cloacae strain). Forty-five patients in a Spanish tertiary care hospital were studied (1996 to 2003). The diversity of In60 and association of In60 with Tn402 or mercury resistance transposons were investigated by overlapping PCR assays and/or hybridization. Plasmid characterization included comparison of restriction fragment length polymorphism patterns and determination of incompatibility group by PCR-based replicon typing, sequencing, and hybridization. CTX-M-9 plasmids belonged to IncHI2 (n = 26), IncP-1α (n = 10), IncFI (n = 4), and IncI (n = 1) groups. Genetic platforms containing blaCTX-M-9 were classified in six types in relation to the In60 backbone and in eight subtypes in relation to Tn402 derivatives. They were associated with Tn21 sequences when located in IncP-1α or IncHI2 plasmids. Our study identified blaCTX-M-9 in a high diversity of CR1-bearing class 1 integrons linked to different Tn402 derivatives, often to Tn21, highlighting the role of recombination events in the evolution of antibiotic resistance plasmids. The presence of blaCTX-M-9 on broad-host-range IncP-1α plasmids might contribute to its dissemination to hosts that were not members of the family Enterobacteriaceae. PMID:16870767

  1. Dengue virus inactivation by minipool TnBP/Triton X-45 treatment of plasma and cryoprecipitate.

    Science.gov (United States)

    Burnouf, T; Chou, M-L; Cheng, L-H; Li, Z-R; Wu, Y-W; El-Ekiaby, M; Tsai, K-H

    2013-01-01

    A minipool solvent/detergent (S/D; 1% TnBP/1% Triton X-45; 31°C) process was developed for viral inactivation of plasma and cryoprecipitate used for transfusion. The goal of this study was to determine the rate and extent of inactivation of dengue virus (DENV) during this process. DENV-1 was propagated using C6/36 mosquito cells to an infectivity titre close to 9 log and spiked (10% v/v) into individual plasma and cryoprecipitate samples from two distinct donors. Samples were taken right after spiking and during viral inactivation treatment by 1% TnBP-1% Triton X-45 at 31°C. DENV-1 infectivity was assessed on Vero E6 cells by a focus-forming assay (FFA). Culture medium and complement-inactivated plasma were used as experimental controls. Experiments were done in duplicate. DENV-1 infectivity was 7·5 log in spiked plasma and 7·1 and 7·3 log in spiked cryoprecipitate. There was no loss of DENV-1 infectivity in the spiked materials, nor in the controls not subjected to S/D treatment. No infectivity was found in plasma and cryoprecipitate subjected to S/D treatment at the first time-point evaluated (10 min). DENV-1 was strongly inactivated in plasma and cryoprecipitate, respectively, within 10 min of 1% TnBP/1% Triton X-45 treatment at 31°C. These data provide a reassurance of the safety of such S/D-treated plasma and cryoprecipitate with regard to the risk of transmission of all DENV serotypes and other flaviviruses. © 2012 The Author(s). Vox Sanguinis © 2012 International Society of Blood Transfusion.

  2. Monoclonal antibodies and cancer

    International Nuclear Information System (INIS)

    Haisma, H.J.

    1987-01-01

    The usefulness of radiolabeled monoclonal antibodies for imaging and treatment of human (ovarian) cancer was investigated. A review of tumor imaging with monoclonal antibodies is presented. Special attention is given to factors that influence the localization of the antibodies in tumors, isotope choice and methods of radiolabeling of the monoclonal antibodies. Two monoclonal antibodies, OC125 and OV-TL3, with high specificity for human epithelial ovarian cancer are characterized. A simple radio-iodination technique was developed for clinical application of the monoclonal antibodies. The behavior of monoclonal antibodies in human tumor xenograft systems and in man are described. Imaging of tumors is complicated because of high background levels of radioactivity in other sites than the tumor, especially in the bloodpool. A technique was developed to improve imaging of human tumor xenographs in nude mice, using subtraction of a specific and a non-specific antibody, radiolabeled with 111 In, 67 Ga and 131 I. To investigate the capability of the two monoclonal antibodies, to specifically localize in human ovarian carcinomas, distribution studies in mice bearing human ovarian carcinoma xenografts were performed. One of the antibodies, OC125, was used for distribution studies in ovarian cancer patients. OC125 was used because of availability and approval to use this antibody in patients. The same antibody was used to investigate the usefulness of radioimmunoimaging in ovarian cancer patients. The interaction of injected radiolabeled antibody OC125 with circulating antigen and an assay to measure the antibody response in ovarian cancer patients after injection of the antibody is described. 265 refs.; 30 figs.; 19 tabs

  3. The TN-RAM - a new cask for shipping high activity irradiated hardware

    International Nuclear Information System (INIS)

    Neider, T.; Hanson, A.S.

    1993-01-01

    Transnuclear, Inc. has developed a Type B(U) radioactive material transport packaging designed specifically for the transport of dry, irradiated, non-fuel bearing components (NFBC). The TN-RAM is a transport cask in the configuration of a right circular cylinder, fabricated from lead and stainless steel, with wood-filled impact limiters attached at both ends. The lead and stainless steel construction of the lid, walls, and bottom provides a shielding effectiveness of approximately 7.1 inches (18 cm) lead equivalent. (J.P.N.)

  4. The (European) HTR Technology Network (HTR-TN) and the development of HTR technology in Europe

    International Nuclear Information System (INIS)

    Hittner, D.; Bogusch, E.; Besson, D.; Verrier, D.; Buckthorpe, D.; Chauvet, V.; Futterer, M.; Van Heek, A.; Lansiart, S.; Phelip, M.; Von Lensa, W.; Pirson, J.

    2007-01-01

    HTR-TN (high temperature reactor - technology network) has been created in 2000 for building a coherent partnership for the development of HTR technology in Europe. For that purpose, HTR-TN elaborated a road-map for the emergence of a new generation of industrial HTRs. Through the fifth EURATOM Framework Programme (FP5), HTR-TN recovered the basis of past European HTR experience, addressed key feasibility issues for Generation IV high temperature systems and made significant advances in the fields of reactor physics (improved calculation methods), fuel (high quality fabrication and very high burn-up behaviour), waste management, qualification of materials for higher performance, component development and definition of safety approach for modular HTRs. In the sixth Framework Programme (FP6), a new integrated project, RAPHAEL, continues the technology developments addressed in FP5 and explores solutions for improving HTR performances towards higher temperatures (above 900 C degrees) and burn-up (up to 200 GWd/tHM): the VHTR objective. Moreover HTR-TN initiated other complementary actions in FP6. The RAPHAEL project has launched key experiments for HTR development: continuation of the graphite irradiation programme started in FP5 in HFR to higher fluences and temperature, test of a heat exchanger element in a helium loop (HE-FUS3, ENEA), irradiation of representative fuel coating material samples for modelling the evolution of their properties, fuel accident heat-up tests in the KUFA facility, integral air ingress tests (NACOK, FZJ), isotopic analysis of fuel irradiated to very high burn-up (170 GWd/tHM). The main motivation for developing a new generation of HTRs is their potential for providing high temperature heat for industrial processes. But coupling a nuclear reactor with an industrial process is very challenging. Therefore after developing base technologies for modular HTRs in FP5 and FP6, the future objective should be the demonstration of such a coupling by

  5. Exploring the Molecular Basis for Selective Binding of Homoserine Dehydrogenase from Mycobacterium leprae TN toward Inhibitors: A Virtual Screening Study

    Directory of Open Access Journals (Sweden)

    Dongling Zhan

    2014-01-01

    Full Text Available Homoserine dehydrogenase (HSD from Mycobacterium leprae TN is an antifungal target for antifungal properties including efficacy against the human pathogen. The 3D structure of HSD has been firmly established by homology modeling methods. Using the template, homoserine dehydrogenase from Thiobacillus denitrificans (PDB Id 3MTJ, a sequence identity of 40% was found and molecular dynamics simulation was used to optimize a reliable structure. The substrate and co-factor-binding regions in HSD were identified. In order to determine the important residues of the substrate (l-aspartate semialdehyde (l-ASA binding, the ASA was docked to the protein; Thr163, Asp198, and Glu192 may be important because they form a hydrogen bond with HSD through AutoDock 4.2 software. neuraminidaseAfter use of a virtual screening technique of HSD, the four top-scoring docking hits all seemed to cation–π ion pair with the key recognition residue Lys107, and Lys207. These ligands therefore seemed to be new chemotypes for HSD. Our results may be helpful for further experimental investigations.

  6. Exploring the molecular basis for selective binding of homoserine dehydrogenase from Mycobacterium leprae TN toward inhibitors: a virtual screening study.

    Science.gov (United States)

    Zhan, Dongling; Wang, Dongmei; Min, Weihong; Han, Weiwei

    2014-01-24

    Homoserine dehydrogenase (HSD) from Mycobacterium leprae TN is an antifungal target for antifungal properties including efficacy against the human pathogen. The 3D structure of HSD has been firmly established by homology modeling methods. Using the template, homoserine dehydrogenase from Thiobacillus denitrificans (PDB Id 3MTJ), a sequence identity of 40% was found and molecular dynamics simulation was used to optimize a reliable structure. The substrate and co-factor-binding regions in HSD were identified. In order to determine the important residues of the substrate (L-aspartate semialdehyde (L-ASA)) binding, the ASA was docked to the protein; Thr163, Asp198, and Glu192 may be important because they form a hydrogen bond with HSD through AutoDock 4.2 software. neuraminidaseAfter use of a virtual screening technique of HSD, the four top-scoring docking hits all seemed to cation-π ion pair with the key recognition residue Lys107, and Lys207. These ligands therefore seemed to be new chemotypes for HSD. Our results may be helpful for further experimental investigations.

  7. [Detection and analysis of anti-Rh blood group antibodies].

    Science.gov (United States)

    Wu, Yuan-jun; Wu, Yong; Chen, Bao-chan; Liu, Yan

    2008-06-01

    To study the prevalence and distribution of anti-Rh blood group antibodies in Chinese population and its clinical significance. Irregular antibodies were screened and identified by Microcolum Gel Coomb's test. For those identified as positive anti-Rh samples, monoclonal antibodies (anti-D, -C, -c, -E and -e) were used to identify the specific antigen and confirm the accuracy of the irregular antibody tests. The titers, Ig-types and 37 Degrees Celsius-reactivity were tested to confirm its clinical significance. For evaluation of the origin of irregular antibodies, histories of pregnancy and transfusion were reviewed. For the newborns who had positive antibodies, their mothers were tested simultaneously to confirm the origin of the antibodies. 47 out of 54 000 (0.087%) patients were identified as positive with Rh blood group antibodies.Of them, 27 cases had history of pregnancy, 13 had transfusion and 1 had the histories of both. 6 newborns had antibodies derived form their mothers. The specificity of the antibody was as follows: 29 with anti-E (61.70%), 8 with anti-D (17.02%), anti-cE 5(10.64%), 4 with anti-c (8.51%) and 1 with anti-C (2.13%). All the 47 Rh blood group antibodies were IgG or IgG+IgM, and were reactive to red blood cells with corresponding antigens at 37 Degrees Celsius, with a highest titer of 1:4 096. The prevalence of Rh antibodies is lower in Chinese population as compared with that in White population.Of all the antibodies, anti-E is most frequently identified and anti-D was declining. Alloimmunization by pregnancy and transfusion is the major cause of Rh antibody production. Rh blood group antibodies derived from mothers are the major cause of Non-ABO-HDN.

  8. The Evaluation of Diagnostic Role of Cardiac Troponin T (cTnT in Newborns with Heart Defects

    Directory of Open Access Journals (Sweden)

    Agata Tarkowska

    2012-01-01

    Full Text Available Heart diseases are a significant cause of morbidity and mortality in newborns. Diagnostic methods are often not sufficient or, in many cases, cannot be used. There is a great advance in medical knowledge concerning biomarkers in the diagnosis of circulatory system in adult patients. Among them, cardiac troponins play the main role. In current literature, there is not enough data concerning the possibility of using them in neonatal cardiac diagnostics. Aim of the Study. To evaluate diagnostic usefulness of cTnT in correlation with other markers of circulatory failure and myocardial damage in newborns with heart defects. Patients and Methods. The study involved 83 newborns up to 46 weeks of postmenstrual age. The exclusion criteria were severe perinatal asphyxia and presence of severe noncardiac diseases. Patients were divided into 2 main groups: group I—54 patients with congenital heart defects (CHDs, and group II (control—29 healthy neonates. All patients underwent detailed examination of circulatory system. Cardiac troponin T (cTnT concentrations were evaluated by Roche CARDIAC T Quantitive test. Results. Performed studies revealed that cTnT levels in newborns with heart pathology were significantly higher than in healthy ones. However, cTnT concentrations in patients with CHD did not correlate with clinical symptoms of heart failure, nor with echocardiographic markers of LV function. Type of heart defect did not influence cTnT levels as well. Only hemodynamic significance evaluated by echocardiography influenced the cTnT levels with statistical significance. Conclusions. (1 Statistically significant differences in cTnT levels between newborns with heart defects and healthy subjects were shown. (2 CTnT levels in newborns with heart defects refer only to hemodynamic significance of the defect.

  9. Radiolabeled antibody imaging

    International Nuclear Information System (INIS)

    Wahl, R.L.

    1987-01-01

    Radiolabeled antibodies, in particular monoclonal antibodies, offer the potential for the specific nuclear imaging of malignant and benign diseases in man. If this imaging potential is realized, they may also have a large role in cancer treatment. This paper reviews: (1) what monoclonal antibodies are and how they differ from polyclonal antibodies, (2) how they are produced and radiolabeled, (3) the results of preclinical and clinical trials in cancer imaging, including the utility of SPECT and antibody fragments, (4) the role of antibodies in the diagnosis of benign diseases, (5) alternate routes of antibody delivery, (6) the role of these agents in therapy, and (7) whether this technology ''revolutionizes'' the practice of nuclear radiology, or has a more limited complementary role in the imaging department

  10. Sensitivity of HIV-1 to neutralization by antibodies against O-linked carbohydrate epitopes despite deletion of O-glycosylation signals in the V3 loop

    DEFF Research Database (Denmark)

    Hansen, J E; Jansson, B; Gram, G J

    1996-01-01

    of such threonine or serine residues could decrease the sensitivity to infectivity inhibition by Tn or sialosyl-Tn specific antibodies. All potentially O-glycosylated threonine and serine residues in the V3 loop of cloned HIV-1 BRU were mutagenized to alanine thus abrogating any O-glycosylation at these sites......It has been suggested that threonine or serine residues in the V3 loop of HIV-1 gp120 are glycosylated with the short-chain O-linked oligosaccharides Tn or sialosyl-Tn that function as epitopes for broadly neutralizing carbohydrate specific antibodies. In this study we examined whether mutation....... Additionally, one of these T-A mutants (T308A) also abrogated the signal for N-glycosylation at N306 inside the V3-loop. The mutant clones were compared with the wild type virus as to sensitivity to neutralization with monoclonal and polyclonal antibodies specific for the tip of the V3 loop of BRU or for the O...

  11. Insertions of transposon Tn5 into ribosomal protein PNA polymerase operons.

    Science.gov (United States)

    Hui, I; Maltman, K; Little, R; Hastrup, S; Johnsen, M; Fiil, N; Dennis, P

    1982-01-01

    The genetic organization and interrelationships between the two ribosomal protein transcription units (the L11 and L10 operons) from near 89 min on the Escherichia coli chromosome were studied by using insertional mutations generated by the kanamycin-resistant transposable element Tn5. The polar effects of Tn5 insertions on the expression of the L11, L1, L10, and L12 ribosomal protein genes and the beta RNA polymerase subunit gene were examined (i) by the level of beta-galactosidase activity generated from L10-lacZ and beta-lacZ gene fusions, (ii) by direct sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the proteins specified by plasmid ribosomal protein genes in UV-irradiated maxicells, and (iii) by urea-polyacrylamide gel electrophoresis of plasmid- and chromosome-specified L12 protein. The results confirmed the organization of these genes into two transcription units as follows: PL11, rplK (L11), rplA (L1), PL10, rplJ (L10), rplL (L12), rpoB (beta). . .; they also localized the position of the PL10 promoter within an 80-nucleotide region near the end of the L1 gene. The results also support the idea that the translational regulatory proteins for the L11 and L10 operons are L1 and L10, respectively, and that the expression of the L12 gene is closely linked to L10 gene expression. Images PMID:6292159

  12. Mini-Tn7 Insertion in an Artificial attTn7 Site Enables Depletion of the Essential Master Regulator CtrA in the Phytopathogen Agrobacterium tumefaciens.

    Science.gov (United States)

    Figueroa-Cuilan, Wanda; Daniel, Jeremy J; Howell, Matthew; Sulaiman, Aliyah; Brown, Pamela J B

    2016-08-15

    Mechanistic studies of many processes in Agrobacterium tumefaciens have been hampered by a lack of genetic tools for characterization of essential genes. In this study, we used a Tn7-based method for inducible control of transcription from an engineered site on the chromosome. We demonstrate that this method enables tighter control of inducible promoters than plasmid-based systems and can be used for depletion studies. The method enables the construction of depletion strains to characterize the roles of essential genes in A. tumefaciens Here, we used the strategy to deplete the alphaproteobacterial master regulator CtrA and found that depletion of this essential gene results in dramatic rounding of cells, which become nonviable. Agrobacterium tumefaciens is a bacterial plant pathogen and natural genetic engineer. Thus, studies of essential processes, including cell cycle progression, DNA replication and segregation, cell growth, and division, may provide insights for limiting disease or improving biotechnology applications. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  13. Monoclonal antibodies to equine CD23 identify the low-affinity receptor for IgE on subpopulations of IgM+ and IgG1+ B-cells in horses.

    Science.gov (United States)

    Wagner, Bettina; Hillegas, Julia M; Babasyan, Susanna

    2012-04-15

    CD23, also called FcεRII, is the low-affinity receptor for IgE and has first been described as a major receptor regulating IgE responses. In addition, CD23 also binds to CD21, integrins and MHC class II molecules and thus has a much wider functional role in immune regulation ranging from involvement in antigen-presentation to multiple cytokine-like functions of soluble CD23. The role of CD23 during immune responses of the horse is less well understood. Here, we expressed equine CD23 in mammalian cells using a novel IL-4 expression system. Expression resulted in high yield of recombinant IL-4/CD23 fusion protein which was purified and used for the generation of monoclonal antibodies (mAbs) to equine CD23. Seven anti-CD23 mAbs were further characterized. The expression of the low-affinity IgE receptor on equine peripheral blood mononuclear cells was analyzed by flow cytometric analysis. Cell surface staining showed that CD23 is mainly expressed by a subpopulation of equine B-cells. Only a very few equine T-cells or monocytes expressed CD23. CD23(+) B-cells were either IgM(+) or IgG1(+) cells. All CD23(+) cells were also positive for cell surface IgE staining suggesting in vivo IgE binding by the receptor. Two of the CD23 mAbs detected either the complete extracellular region of CD23 or a 22kDa cleavage product of CD23 by Western blotting. The new anti-CD23 mAbs provide valuable reagents to further analyze the roles of CD23 during immune responses of the horse in health and disease. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. Development of an in Vitro Potency Assay for Anti-anthrax Lethal Toxin Neutralizing Antibodies

    Directory of Open Access Journals (Sweden)

    Sjoerd Rijpkema

    2012-01-01

    Full Text Available Lethal toxin (LT of Bacillus anthracis reduces the production of a number of inflammatory mediators, including transcription factors, chemokines and cytokines in various human cell lines, leading to down-regulation of the host inflammatory response. Previously we showed that the reduction of interleukin-8 (IL-8 is a sensitive marker of LT-mediated intoxication in human neutrophil-like NB-4 cells and that IL-8 levels are restored to normality when therapeutic monoclonal antibodies (mAb with toxin-neutralising (TN activity are added. We used this information to develop cell-based assays that examine the effects of TN therapeutic mAbs designed to treat LT intoxication and here we extend these findings. We present an in vitro assay based on human endothelial cell line HUVEC jr2, which measures the TN activity of therapeutic anti-LT mAbs using IL-8 as a marker for intoxication. HUVEC jr2 cells have the advantage over NB-4 cells that they are adherent, do not require a differentiation step and can be used in a microtitre plate format and therefore can facilitate high throughput analysis. This human cell-based assay provides a valid alternative to the mouse macrophage assay as it is a more biologically relevant model of the effects of toxin-neutralising antibodies in human infection.

  15. Integrative Gene Cloning and Expression System for Streptomyces sp. US 24 and Streptomyces sp. TN 58 Bioactive Molecule Producing Strains

    Directory of Open Access Journals (Sweden)

    Samiha Sioud

    2009-01-01

    Full Text Available Streptomyces sp. US 24 and Streptomyces sp. TN 58, two strains producing interesting bioactive molecules, were successfully transformed using E. coli ET12567 (pUZ8002, as a conjugal donor, carrying the integrative plasmid pSET152. For the Streptomyces sp. US 24 strain, two copies of this plasmid were tandemly integrated in the chromosome, whereas for Streptomyces sp. TN 58, the integration was in single copy at the attB site. Plasmid pSET152 was inherited every time for all analysed Streptomyces sp. US 24 and Streptomyces sp. TN 58 exconjugants under nonselective conditions. The growth, morphological differentiation, and active molecules production of all studied pSET152 integrated exconjugants were identical to those of wild type strains. Consequently, conjugal transfer using pSET152 integration system is a suitable means of genes transfer and expression for both studied strains. To validate the above gene transfer system, the glucose isomerase gene (xylA from Streptomyces sp. SK was expressed in strain Streptomyces sp. TN 58. Obtained results indicated that heterologous glucose isomerase could be expressed and folded effectively. Glucose isomerase activity of the constructed TN 58 recombinant strain is of about eighteenfold higher than that of the Streptomyces sp. SK strain. Such results are certainly of importance due to the potential use of improved strains in biotechnological process for the production of high-fructose syrup from starch.

  16. Structural Analysis of the Hg(II)-Regulatory Protein Tn501 MerR from Pseudomonas aeruginosa

    Science.gov (United States)

    Wang, Dan; Huang, Shanqing; Liu, Pingying; Liu, Xichun; He, Yafeng; Chen, Weizhong; Hu, Qingyuan; Wei, Tianbiao; Gan, Jianhua; Ma, Jing; Chen, Hao

    2016-09-01

    The metalloprotein MerR is a mercury(II)-dependent transcriptional repressor-activator that responds to mercury(II) with extraordinary sensitivity and selectivity. It’s widely distributed in both Gram-negative and Gram-positive bacteria but with barely detectable sequence identities between the two sources. To provide structural basis for the considerable biochemical and biophysical experiments previously performed on Tn501 and Tn21 MerR from Gram-negative bacteria, we analyzed the crystal structure of mercury(II)-bound Tn501 MerR. The structure in the metal-binding domain provides Tn501 MerR with a high affinity for mercury(II) and the ability to distinguish mercury(II) from other metals with its unique planar trigonal coordination geometry, which is adopted by both Gram-negative and Gram-positive bacteria. The mercury(II) coordination state in the C-terminal metal-binding domain is transmitted through the allosteric network across the dimer interface to the N-terminal DNA-binding domain. Together with the previous mutagenesis analyses, the present data indicate that the residues in the allosteric pathway have a central role in maintaining the functions of Tn501 MerR. In addition, the complex structure exhibits significant differences in tertiary and quaternary structural arrangements compared to those of Bacillus MerR from Gram-positive bacteria, which probably enable them to function with specific promoter DNA with different spacers between -35 and -10 elements.

  17. Prevalence of aac(6')-Ie-aph(2″)-Ia resistance gene and its linkage to Tn5281 in Enterococcus faecalis and Enterococcus faecium isolates from Tabriz hospitals.

    Science.gov (United States)

    Behnood, Amir; Farajnia, Safar; Moaddab, Seyed Reza; Ahdi-Khosroshahi, Shiva; Katayounzadeh, Aliakbar

    2013-09-01

    High-level gentamicin resistance (HLGR: MIC ≥ 500 µg/ml) in Enterococci is mediated by aminoglycoside modifying enzymes which is mainly encoded by aac(6')-Ie-aph(2″)-Ia gene. The aim of this study was to evaluate the frequency of aac(6')-Ie-aph(2″)-Ia gene in clinical isolates of Enterococcus facium and Enterococcus faecalis collected from hospitals in northwest of Iran. In the present study a total of 111 enterococcus isolates were collected from 4 hospitals during a two year period (July 2009-August 2011). Bacterial identification and species determination were carried out by standard biochemical tests. Antimicrobial susceptibility was evaluated by Kirby Bauer disc diffusion method. MICs were determined by agar dilution method. The frequency of aac(6')-Ie-aph(2″)-Ia gene in the isolates was determined by PCR. The carriage of resistance gene on Tn5281 transposon was identified by long PCR and dot-blot hybridization methods. Antibiotic susceptibility tests revealed that the highest resistance was against streptomycin (74.77%) and erythromycin (67.58%) whereas the highest susceptibility was observed to vancomycin (81.1%). 36 isolates (32.43%) were identified as HLGR, 34(94.44%) of them had resistant gene in their genome. Long PCR studies revealed that 88% of HLGR clinical isolates harboured Tn5281. The aac(6')-Ie-aph(2″)-Ia resistance gene was present on Tn5281 transposon in all 32 isolates according to dot blot hybridization test. The results of this study indicated that aac(6')-Ie-aph(2″)-Ia resistance gene is highly prevalent in gentamicin resistant isolates. Carriage of aac(6')-Ie-aph(2″)-Ia resistance gene on Tn5281 transposable element suggests possible contribution of this transposone on dissemination of resistance gene among enterococcus isolates.

  18. Serum Antibody Biomarkers for ASD

    Science.gov (United States)

    2015-10-01

    45-56. Singh VK. (2009) Phenotypic expression of autoimmune autistic disorder (AAD): A major subset of autism. Ann Clin Psychiat. 21:148-160. 5...spectrum disorder (ASD) is a neurodevelopmental disorder characterized by impairments in communication (verbal and nonverbal), social interactions, and... autoimmunity ; in particular, the generation of antibodies reactive against brain and CNS proteins. The goal of this grant is to identify serum

  19. Transitioning high sensitivity cardiac troponin I (hs-cTnI) into routine diagnostic use: More than just a sensitivity issue

    LENUS (Irish Health Repository)

    Lee, Graham R

    2016-04-01

    High sensitivity cardiac troponin T and I (hs-cTnT and hs-cTnI) assays show analytical, diagnostic and prognostic improvement over contemporary sensitive cTn assays. However, given the importance of troponin in the diagnosis of myocardial infarction, implementing this test requires rigorous analytical and clinical verification across the total testing pathway. This was the aim of this study.

  20. Engineered CAR T Cells Targeting the Cancer-Associated Tn-Glycoform of the Membrane Mucin MUC1 Control Adenocarcinoma

    DEFF Research Database (Denmark)

    Posey, Avery D; Schwab, Robert D; Boesteanu, Alina C

    2016-01-01

    Genetically modified T cells expressing chimeric antigen receptors (CARs) demonstrate robust responses against lineage restricted, non-essential targets in hematologic cancers. However, in solid tumors, the full potential of CAR T cell therapy is limited by the availability of cell surface antigens......-associated Tn glycoform of MUC1, a neoantigen expressed in a variety of cancers. Anti-Tn-MUC1 CAR T cells demonstrated target-specific cytotoxicity and successfully controlled tumor growth in xenograft models of T cell leukemia and pancreatic cancer. These findings demonstrate the therapeutic efficacy of CAR T...... cells directed against Tn-MUC1 and present aberrantly glycosylated antigens as a novel class of targets for tumor therapy with engineered T cells....

  1. Development Of Protocols For Simultaneous Measurements Of Rn, Tn Using Nuclear Track Detectors And Trial Application In Mining Areas Of Vietnam

    International Nuclear Information System (INIS)

    Bui Dac Dung; Trinh Van Giap; Le Dinh Cuong; Tran Khanh Minh; Nguyen Huu Quyet; Nguyen Van Khanh; Tran Ngoc Toan

    2013-01-01

    Radioactive gases Radon (Rn) and Thoron (Tn) contribute of more than 50% of natural radiation dose. However, separate measurements of Rn and Tn have not been paid enough attention. An Intergovernmental Cooperation Project was conducted at the Institute for Nuclear Science and Technology with the help from Hungarian experts. The main tasks of the project were to finalize 02 protocols for simultaneous measurements of Rn and Tn using nuclear track detectors and to test these protocols in investigating the concentrations of Rn and Tn, calculating the natural dose due to Rn and Tn, and evaluating the increased radiation dose and radiation safety due to mining activities. Main results of the project include 02 protocols for simultaneous measurements of Rn and Tn and the test results in the mining areas. Rn and Tn concentrations inside the coal mining tunnels are in the average level of Vietnam and the world. Tn concentration inside the factory for separating Zircon at the Ha Tinh Zircon Processing Plant was found to be very high, up to 2931 Bq/m 3 . Based on annual effective dose calculation, workers inside the factory for separating Zircon at the Ha Tinh Zircon Processing Plant could receive an annual effective dose due to Rn and Tn of 4.890 mSv/year, and the increasing dose of 4.710 mSv/year is for higher than 1 mSv/year recommended by the IAEA. (author)

  2. Challenges in harmonizing integrated healthcare network laboratories: Multi-center evaluation of the AccuTnI+3 troponin assay.

    Science.gov (United States)

    Greene, Dina N; Holmes, Daniel T; Liang, Joy; Kwong, Shiu-Land; Lorey, Thomas S; Petrie, Matthew S

    2015-03-01

    Beckman Coulter has recently introduced a new troponin assay manufactured for the Access2 and DxI platforms, releasing it under the name AccuTnI+3. Clinical laboratories are required to validate method performance before testing and reporting patient results. Beckman Coulter Access 2 instruments (n=42) across Kaiser Permanente Northern California were evaluated for their performance characteristics using the AccuTnI+3 reagent. Precision, linearity, and patient sample comparisons were performed on each instrument. Limit of the blank (LOB), limit of detection (LOD), limit of quantitation (LOQ), serum plasma comparisons, and specimen stability were evaluated using a single instrument. The assay was linear from 0-100,000ng/L. The LOB, LOD and LOQ were determined to be 5, 8 and 20ng/L, respectively. Interday precision on the low QC (mean concentration 41ng/L) ranged from 3.0% to 14.2%. The bias observed between the former assay (AccuTnI) and the AccuTnI+3 was comparable to the inter-instrument bias for either assay. Non-uniform distribution was observed in the precision and inter-instrument/inter-assay comparisons among the instruments evaluated. The AccuTnI and AccuTnI+3 troponin assays are equivalent across the analytical measuring range. There was no significant difference at the medical decision point. No changes in patient results are anticipated. However, the assay-independent inter-instrument bias observed is an important consideration for harmonization efforts. Copyright © 2014. Published by Elsevier Inc.

  3. Completion of the nucleotide sequence of the central region of Tn5 confirms the presence of three resistance genes.

    OpenAIRE

    Mazodier, P; Cossart, P; Giraud, E; Gasser, F

    1985-01-01

    The DNA sequence of the region located downstream from the kanamycin resistance gene of Tn5 up to the right inverted repeat IS50R has been determined. This completes the determination of the sequence of Tn5 which is 5818 bp long. The 2.7 Kb central region contains three resistance genes: the kanamycin-neomycin resistance gene, a gene coding for resistance to CL990 an antimitotic-antibiotic compound of the bleomycin family and a third gene that confers streptomycin resistance in some bacterial...

  4. Tethered-variable CL bispecific IgG: an antibody platform for rapid bispecific antibody screening.

    Science.gov (United States)

    Kim, Hok Seon; Dunshee, Diana Ronai; Yee, Angie; Tong, Raymond K; Kim, Ingrid; Farahi, Farzam; Hongo, Jo-Anne; Ernst, James A; Sonoda, Junichiro; Spiess, Christoph

    2017-09-01

    Bispecific antibodies offer a clinically validated platform for drug discovery. In generating functionally active bispecific antibodies, it is necessary to identify a unique parental antibody pair to merge into a single molecule. However, technologies that allow high-throughput production of bispecific immunoglobulin Gs (BsIgGs) for screening purposes are limited. Here, we describe a novel bispecific antibody format termed tethered-variable CLBsIgG (tcBsIgG) that allows robust production of intact BsIgG in a single cell line, concurrently ensuring cognate light chain pairing and preserving key antibody structural and functional properties. This technology is broadly applicable in the generation of BsIgG from a variety of antibody isotypes, including human BsIgG1, BsIgG2 and BsIgG4. The practicality of the tcBsIgG platform is demonstrated by screening BsIgGs generated from FGF21-mimetic anti-Klotho-β agonistic antibodies in a combinatorial manner. This screen identified multiple biepitopic combinations with enhanced agonistic activity relative to the parental monoclonal antibodies, thereby demonstrating that biepitopic antibodies can acquire enhanced functionality compared to monospecific parental antibodies. By design, the tcBsIgG format is amenable to high-throughput production of large panels of bispecific antibodies and thus can facilitate the identification of rare BsIgG combinations to enable the discovery of molecules with improved biological function. © The Author 2017. Published by Oxford University Press.

  5. Tn5-induced pBS286 plasmid mutations blocking early stages of napthalene oxidation

    International Nuclear Information System (INIS)

    Kosheleva, I.A.; Tsoi, T.V.; Ivashina, T.V.; Selifonov, S.A.; Starovoitov, I.I.; Boronin, A.M.

    1988-01-01

    The authors present data on the further analysis of the structural and functional organization of the nah region of plasmid pBS286 controlling the constitutive oxidation of naphthalene by Pseudomonas putida cells. They have studied Tn5-induced mutations blocking early stages of naphthalene oxidation. They present and discuss data providing evidence that, in contrast to plasmid NAH7, the mechanism of regulation of the nahl operon of plasmid NPL-1, the parent plasmid of plasmid pBS286, with inducible synthesis of naphthalene dioxygenase can include elements of a negative control with participation of the regulatory locus R, located proximal to the structural nah genes and closely linked to or overlapped by the inverted control DNA segment (4.2 kb). They also present data on the possibility of regulation of the activity of the catechol-splitting meta-pathway genes with the participation of products of early stages of naphthalene oxidation

  6. Loading and transport of high-active waste (HAW) with the TN85 flask in 2008

    Energy Technology Data Exchange (ETDEWEB)

    Rys, Michael; Horn, Thomas; Graf, Wilhelm [GNS Gesellschaft fuer Nuklear-Service mbH (Germany); Bonface, Jean-Michael [TN International, Montigny-le-Bretonneux (France)

    2009-07-01

    As a part of the operation of nuclear power plants, it is essential to safely manage the radioactive waste. With new developments in science and technology, it is a dynamic process to adapt procedures, equipment and flasks to be used in the future. This is a task for specialists - a task for GNS Gesellschaft fuer Nuklear-Service mbH and for TN International. Until 1994 reprocessing of spent fuel from German nuclear power plants was mandatory for the Utilities (EVU) in Germany. Basis for the reprocessing was the German Atomic Act. The German Utilities concluded contracts on reprocessing with Compagnie Generale des Matieres Nucleaires (COGEMA, now AREVA NC) in France and British Nuclear Fuels plc (BNFL, now INS) in England. The total amount to be reprocessed comes to 5309 t HM contracted to AREVA NC and 768 t HM contracted to INS. The waste generated from reprocessing - or an equivalent amount of radioactive material - has to be returned to the country of origin. In 1979 already an exchange of notes took place between the German and the French government with the obligation of both sides to enable and support the return of reprocessing residues or equivalents. The return of high-active waste (HAW) from France has started in 1996 with the first attribution of 28 glass canisters (one flask) to German Utilities by AREVA NC. Until 2007, 75 flasks loaded with vitrified residue (VR) canisters have been transported to Gorleben. For these transports CASTOR {sup registered} HAW 20/28 CG flasks have been used. This presentation will give some background information about the last HAW transport in 2008 with the new flask generation of the type TN85. It will also describe the assembly of the new flask, the preparation of the flask for the loading campaign as well as the loading procedure. (orig.)

  7. TN-68 Spent Fuel Transport Cask Analytical Evaluation for Drop Events

    International Nuclear Information System (INIS)

    Shah, M.J.; Klymyshyn, Nicholas A.; Adkins, Harold E.; Koeppel, Brian J.

    2007-01-01

    The U.S. Nuclear Regulatory Commission (NRC) is responsible for licensing commercial spent nuclear fuel transported in casks certified by NRC under the Code of Federal Regulations (10 CFR), Title 10, Part 71 (1). Both the International Atomic Energy Agency regulations for transporting radioactive materials (2, paragraph 727), and 10 CFR 71.73 require casks to be evaluated for hypothetical accident conditions, which includes a 9-meter (m) (30-ft) drop-impact event onto a flat, essentially unyielding, horizontal surface, in the most damaging orientation. This paper examines the behavior of one of the NRC certified transportation casks, the TN-68 (3), for drop-impact events. The specific area examined is the behavior of the bolted connections in the cask body and the closure lid, which are significantly loaded during the hypothetical drop-impact event. Analytical work to evaluate the NRC-certified TN-68 spent fuel transport cask (3) for a 9-m (30-ft) drop-impact event on a flat, unyielding, horizontal surface, was performed using the ANSYS (4) and LS DYNA (5) finite-element analysis codes. The models were sufficiently detailed, in the areas of bolt closure interfaces and containment boundaries, to evaluate the structural integrity of the bolted connections under 9-m (30-ft) free-drop hypothetical accident conditions, as specified in 10 CFR 71.73. Evaluation of the cask for puncture, caused by a free drop through a distance of 1-m (40-in.) onto a mild steel bar mounted on a flat, essentially unyielding, horizontal surface, required by 10 CFR 71.73, was not included in the current work, and will have to be addressed in the future. Based on the analyses performed to date, it is concluded that, even though brief separation of the flange and the lid surfaces may occur under some conditions, the seals would close at the end of the drop events, because the materials remain elastic during the duration of the event

  8. Antibodies Against Melanin

    African Journals Online (AJOL)

    1973-01-06

    Jan 6, 1973 ... Departments of Internal Medicine and Anatomical Pathology, University of Stellenbosch and MRC. Pigment Metabolism Research Unit, ... at the production of antibodies against natural melanoprotein. and a consideration of our negative .... the random polymerization of several monomers, antibody formed ...

  9. Recombinant renewable polyclonal antibodies.

    Science.gov (United States)

    Ferrara, Fortunato; D'Angelo, Sara; Gaiotto, Tiziano; Naranjo, Leslie; Tian, Hongzhao; Gräslund, Susanne; Dobrovetsky, Elena; Hraber, Peter; Lund-Johansen, Fridtjof; Saragozza, Silvia; Sblattero, Daniele; Kiss, Csaba; Bradbury, Andrew R M

    2015-01-01

    Only a small fraction of the antibodies in a traditional polyclonal antibody mixture recognize the target of interest, frequently resulting in undesirable polyreactivity. Here, we show that high-quality recombinant polyclonals, in which hundreds of different antibodies are all directed toward a target of interest, can be easily generated in vitro by combining phage and yeast display. We show that, unlike traditional polyclonals, which are limited resources, recombinant polyclonal antibodies can be amplified over one hundred million-fold without losing representation or functionality. Our protocol was tested on 9 different targets to demonstrate how the strategy allows the selective amplification of antibodies directed toward desirable target specific epitopes, such as those found in one protein but not a closely related one, and the elimination of antibodies recognizing common epitopes, without significant loss of diversity. These recombinant renewable polyclonal antibodies are usable in different assays, and can be generated in high throughput. This approach could potentially be used to develop highly specific recombinant renewable antibodies against all human gene products.

  10. Tn7::In2-8 dispersion in multidrug resistant isolates of Acinetobacter baumannii from Chile Dispersión de Tn7::In2-8 en aislamientos multirresistentes de Acinetobacter baumannii de Chile

    Directory of Open Access Journals (Sweden)

    M. S. Ramírez

    2010-06-01

    Full Text Available Acinetobacter baumannii is considered an important pathogen in our hospital environment having a well-known capacity to acquire different mechanisms of antibiotic resistance. Previous studies in our laboratory had exposed the high dispersion of class 2 integrons in this species. In the present study, we analyzed 7 multiresistant intI2 positive A. baumannii isolates, 6 of which were found to harbour the Tn7::In2-8 element. Our results demonstrate the unusually high distribution of Tn7::In2-8 among different A. baumannii clones from Chile, suggesting a particular behavior of these elements at geographical level.Acinetobacter baumannii, patógeno de importancia clínica en el ámbito hospitalario, es reconocido como un microorganismo que posee la capacidad de evolucionar rápidamente hacia la multirresistencia. Estudios previos efectuados en nuestro laboratorio han demostrado la alta dispersión de los integrones de clase 2 en aislamientos de esta especie. En el presente trabajo se analizaron 7 aislamientos de Acinetobacter baumannii multirresistentes portadores de la integrasa de clase 2, 6 de los cuales portaban el inusual arreglo Tn7::In2-8. Nuestros resultados muestran una elevada frecuencia de dispersión del elemento Tn7::In2-8 en diferentes clones circulantes en Chile, lo que sugiere un comportamiento geográfico particular.

  11. Identification and Characterization of Non-Cellulose-Producing Mutants of Gluconacetobacter hansenii Generated by Tn5 Transposon Mutagenesis

    Science.gov (United States)

    Deng, Ying; Nagachar, Nivedita; Xiao, Chaowen; Tien, Ming

    2013-01-01

    The acs operon of Gluconacetobacter is thought to encode AcsA, AcsB, AcsC, and AcsD proteins that constitute the cellulose synthase complex, required for the synthesis and secretion of crystalline cellulose microfibrils. A few other genes have been shown to be involved in this process, but their precise role is unclear. We report here the use of Tn5 transposon insertion mutagenesis to identify and characterize six non-cellulose-producing (Cel−) mutants of Gluconacetobacter hansenii ATCC 23769. The genes disrupted were acsA, acsC, ccpAx (encoding cellulose-complementing protein [the subscript “Ax” indicates genes from organisms formerly classified as Acetobacter xylinum]), dgc1 (encoding guanylate dicyclase), and crp-fnr (encoding a cyclic AMP receptor protein/fumarate nitrate reductase transcriptional regulator). Protein blot analysis revealed that (i) AcsB and AcsC were absent in the acsA mutant, (ii) the levels of AcsB and AcsC were significantly reduced in the ccpAx mutant, and (iii) the level of AcsD was not affected in any of the Cel− mutants. Promoter analysis showed that the acs operon does not include acsD, unlike the organization of the acs operon of several strains of closely related Gluconacetobacter xylinus. Complementation experiments confirmed that the gene disrupted in each Cel− mutant was responsible for the phenotype. Quantitative real-time PCR and protein blotting results suggest that the transcription of bglAx (encoding β-glucosidase and located immediately downstream from acsD) was strongly dependent on Crp/Fnr. A bglAx knockout mutant, generated via homologous recombination, produced only ∼16% of the wild-type cellulose level. Since the crp-fnr mutant did not produce any cellulose, Crp/Fnr may regulate the expression of other gene(s) involved in cellulose biosynthesis. PMID:24013627

  12. Expression of sialyl-Tn sugar antigen in bladder cancer cells affects response toBacillus Calmette Guérin(BCG) and to oxidative damage.

    Science.gov (United States)

    Severino, Paulo F; Silva, Mariana; Carrascal, Mylene; Malagolini, Nadia; Chiricolo, Mariella; Venturi, Giulia; Astolfi, Annalisa; Catera, Mariangela; Videira, Paula A; Dall'Olio, Fabio

    2017-08-15

    The sialyl-Tn (sTn) antigen is an O -linked carbohydrate chain aberrantly expressed in bladder cancer (BC), whose biosynthesis is mainly controlled by the sialyltransferase ST6GALNAC1. Treatment with Bacillus Calmette-Guérin (BCG) is the most effective adjuvant immunotherapy for superficial BC but one third of the patients fail to respond. A poorly understood correlation between the expression of sTn and BC patient's response to BCG was previously observed. By analyzing tumor tissues, we showed that patients with high ST6GALNAC1 and IL-6 mRNA expression were BCG responders. To investigate the role of sTn in BC cell biology and BCG response, we established the cell lines MCR sTn and MCR Nc by retroviral transduction of the BC cell line MCR with the ST6GALNAC1 cDNA or with an empty vector, respectively. Compared with MCR Nc , BCG-stimulated MCR sTn secreted higher levels of IL-6 and IL-8 and their secretome induced a stronger IL-6, IL-1β, and TNFα secretion by macrophages, suggesting the induction of a stronger inflammatory response. Transcriptomic analysis of MCR Nc and MCR sTn revealed that ST6GALNAC1 /sTn expression modulates hundreds of genes towards a putative more malignant phenotype and down-regulates several genes maintaining genomic stability. Consistently, MCR sTn cells displayed higher H 2 O 2 sensitivity. In MCR sTn ,, BCG challenge induced an increased expression of several regulatory non coding RNA genes. These results indicate that the expression of ST6GALNAC1 /sTn improves the response to BCG therapy by inducing a stronger macrophage response and alters gene expression towards malignancy and genomic instability, increasing the sensitivity of BC cells to the oxidizing agents released by BCG.

  13. Structure Based Antibody-Like Peptidomimetics

    Directory of Open Access Journals (Sweden)

    Mark I. Greene

    2012-02-01

    Full Text Available Biologics such as monoclonal antibodies (mAb and soluble receptors represent new classes of therapeutic agents for treatment of several diseases. High affinity and high specificity biologics can be utilized for variety of clinical purposes. Monoclonal antibodies have been used as diagnostic agents when coupled with radionuclide, immune modulatory agents or in the treatment of cancers. Among other limitations of using large molecules for therapy the actual cost of biologics has become an issue. There is an effort among chemists and biologists to reduce the size of biologics which includes monoclonal antibodies and receptors without a reduction of biological efficacy. Single chain antibody, camel antibodies, Fv fragments are examples of this type of deconstructive process. Small high-affinity peptides have been identified using phage screening. Our laboratory used a structure-based approach to develop small-size peptidomimetics from the three-dimensional structure of proteins with immunoglobulin folds as exemplified by CD4 and antibodies. Peptides derived either from the receptor or their cognate ligand mimics the functions of the parental macromolecule. These constrained peptides not only provide a platform for developing small molecule drugs, but also provide insight into the atomic features of protein-protein interactions. A general overview of the reduction of monoclonal antibodies to small exocyclic peptide and its prospects as a useful diagnostic and as a drug in the treatment of cancer are discussed.

  14. Antibody engineering: methods and protocols

    National Research Council Canada - National Science Library

    Chames, Patrick

    2012-01-01

    "Antibody Engineering: Methods and Protocols, Second Edition was compiled to give complete and easy access to a variety of antibody engineering techniques, starting from the creation of antibody repertoires and efficient...

  15. Anti-insulin antibody test

    Science.gov (United States)

    Insulin antibodies - serum; Insulin Ab test; Insulin resistance - insulin antibodies; Diabetes - insulin antibodies ... You appear to have an allergic response to insulin Insulin no longer seems to control your diabetes

  16. Survivors Remorse: antibody-mediated protection against HIV-1.

    Science.gov (United States)

    Lewis, George K; Pazgier, Marzena; DeVico, Anthony L

    2017-01-01

    It is clear that antibodies can play a pivotal role in preventing the transmission of HIV-1 and large efforts to identify an effective antibody-based vaccine to quell the epidemic. Shortly after HIV-1 was discovered as the cause of AIDS, the search for epitopes recognized by neutralizing antibodies became the driving strategy for an antibody-based vaccine. Neutralization escape variants were discovered shortly thereafter, and, after almost three decades of investigation, it is now known that autologous neutralizing antibody responses and their selection of neutralization resistant HIV-1 variants can lead to broadly neutralizing antibodies in some infected individuals. This observation drives an intensive effort to identify a vaccine to elicit broadly neutralizing antibodies. In contrast, there has been less systematic study of antibody specificities that must rely mainly or exclusively on other protective mechanisms, although non-human primate (NHP) studies as well as the RV144 vaccine trial indicate that non-neutralizing antibodies can contribute to protection. Here we propose a novel strategy to identify new epitope targets recognized by these antibodies for which viral escape is unlikely or impossible. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  17. Assessing Tn5 and Sleeping Beauty for transpositional transgenesis by cytoplasmic injection into bovine and ovine zygotes

    Science.gov (United States)

    Bevacqua, R. J.; Fernandez-Martin, R.; Canel, N. G.; Gibbons, A.; Texeira, D.; Lange, F.; Vans Landschoot, G.; Savy, V.; Briski, O.; Hiriart, M. I.; Grueso, E.; Ivics, Z.; Taboga, O.; Kues, W. A.; Ferraris, S.

    2017-01-01

    Transgenic domestic animals represent an alternative to bioreactors for large-scale production of biopharmaceuticals and could also provide more accurate biomedical models than rodents. However, their generation remains inefficient. Recently, DNA transposons allowed improved transgenesis efficiencies in mice and pigs. In this work, Tn5 and Sleeping Beauty (SB) transposon systems were evaluated for transgenesis by simple cytoplasmic injection in livestock zygotes. In the case of Tn5, the transposome complex of transposon nucleic acid and Tn5 protein was injected. In the case of SB, the supercoiled plasmids encoding a transposon and the SB transposase were co-injected. In vitro produced bovine zygotes were used to establish the cytoplasmic injection conditions. The in vitro cultured blastocysts were evaluated for reporter gene expression and genotyped. Subsequently, both transposon systems were injected in seasonally available ovine zygotes, employing transposons carrying the recombinant human factor IX driven by the beta-lactoglobulin promoter. The Tn5 approach did not result in transgenic lambs. In contrast, the Sleeping Beauty injection resulted in 2 lambs (29%) carrying the transgene. Both animals exhibited cellular mosaicism of the transgene. The extraembryonic tissues (placenta or umbilical cord) of three additional animals were also transgenic. These results show that transpositional transgenesis by cytoplasmic injection of SB transposon components can be applied for the production of transgenic lambs of pharmaceutical interest. PMID:28301581

  18. 78 FR 21099 - Grant of Authority for Subzone Status, Hemlock Semiconductor, L.L.C., (Polysilicon), Clarksville, TN

    Science.gov (United States)

    2013-04-09

    ... Status, Hemlock Semiconductor, L.L.C., (Polysilicon), Clarksville, TN Pursuant to its authority under the... manufacturing authority at the polysilicon manufacturing facility of Hemlock Semiconductor, L.L.C., located in... status for activity related to the manufacturing of polysilicon at the facility of Hemlock Semiconductor...

  19. 77 FR 73978 - Foreign-Trade Zone 148-Knoxville, TN, Toho Tenax America, Inc. (Carbon Fiber Manufacturing...

    Science.gov (United States)

    2012-12-12

    ... manufacture carbon fiber for export and oxidized polyacrylonitrile fiber (Board Order 1868, 77 FR 69435, 11/19/2012). Board Order 1868 did not include authority to manufacture carbon fiber for the U.S. market; the...--Knoxville, TN, Toho Tenax America, Inc. (Carbon Fiber Manufacturing Authority), Opening of Comment Period on...

  20. 76 FR 34799 - Permanent Dam Safety Modification at Cherokee, Fort Loudoun, Tellico, and Watts Bar Dams, TN

    Science.gov (United States)

    2011-06-14

    ... TENNESSEE VALLEY AUTHORITY Permanent Dam Safety Modification at Cherokee, Fort Loudoun, Tellico, and Watts Bar Dams, TN AGENCY: Tennessee Valley Authority. ACTION: Notice of intent. SUMMARY: This... existing dam facilities at Cherokee, Fort Loudoun, Tellico, and Watts Bar dams in Tennessee. The level of...

  1. 77 FR 21505 - Proposed Amendment of Class D and E Airspace; Blountville, TN, and Revocation of Class E Airspace...

    Science.gov (United States)

    2012-04-10

    ... Airport, VA (Lat. 36[deg]41'14'' N., long. 82[deg]02'00'' W.) Rogersville, Hawkins County Airport, TN (Lat... Highlands Airport and within a 7-mile radius of Hawkins County Airport, and within 7 miles each side of Runway 07/25 centerline, extending from the 7-mile radius to 12 miles east of Hawkins County Airport...

  2. 75 FR 38979 - Expansion/Reorganization of Foreign-Trade Zone 204, Tri-Cities Area, TN/VA

    Science.gov (United States)

    2010-07-07

    ... DEPARTMENT OF COMMERCE Foreign-Trade Zones Board [Order No. 1691] Expansion/Reorganization of Foreign-Trade Zone 204, Tri-Cities Area, TN/VA Pursuant to its authority under the Foreign-Trade Zones Act of June 18, 1934, as amended (19 U.S.C. 81a-81u), the Foreign-Trade Zones Board (the Board) adopts...

  3. Highly specific PET imaging of prostate tumors in mice with an iodine-124-labeled antibody fragment that targets phosphatidylserine.

    Directory of Open Access Journals (Sweden)

    Jason H Stafford

    Full Text Available Phosphatidylserine (PS is an attractive target for imaging agents that identify tumors and assess their response to therapy. PS is absent from the surface of most cell types, but becomes exposed on tumor cells and tumor vasculature in response to oxidative stresses in the tumor microenvironment and increases in response to therapy. To image exposed PS, we used a fully human PS-targeting antibody fragment, PGN635 F(ab'2, that binds to complexes of PS and β2-glycoprotein I. PGN635 F(ab'2 was labeled with the positron-emitting isotope iodine-124 ((124I and the resulting probe was injected into nude mice bearing subcutaneous or orthotopic human PC3 prostate tumors. Biodistribution studies showed that (124I-PGN635 F(ab'2 localized with remarkable specificity to the tumors with little uptake in other organs, including the liver and kidneys. Clear delineation of the tumors was achieved by PET 48 hours after injection. Radiation of the tumors with 15 Gy or systemic treatment of the mice with 10 mg/kg docetaxel increased localization in the tumors. Tumor-to-normal (T/N ratios were inversely correlated with tumor growth measured over 28 days. These data indicate that (124I-PGN635 F(ab'2 is a promising new imaging agent for predicting tumor response to therapy.

  4. The effect and biological mechanism of COD/TN ratio on nitrogen removal in a novel upflow microaerobic sludge reactor treating manure-free piggery wastewater.

    Science.gov (United States)

    Li, Jianzheng; Meng, Jia; Li, Jiuling; Wang, Cheng; Deng, Kaiwen; Sun, Kai; Buelna, Gerardo

    2016-06-01

    A novel upflow microaerobic sludge reactor (UMSR) was constructed to treat manure-free piggery wastewater with high NH4(+)-N concentration and low COD/TN ratio, and the effect and biological mechanism of COD/TN ratio on nitrogen removal were investigated at a constant hydraulic retention time of 8h and 35°C. The results showed that the UMSR could treat the wastewater with a better synchronous removal of COD, NH4(+)-N and TN. The microaerobic UMSR allowed nitrifiers, and heterotrophic and autotrophic denitrifiers to thrive in the flocs, revealing a multiple nitrogen removal mechanism in the reactor. Both the nitrifiers and denitrifiers would be restricted by an influent COD/TN ratio more than 0.82, resulting in a decrease of TN removal in the UMSR. To get a TN removal over 80% with a TN load removal above 0.86kg/(m(3)·d) in the UMSR, the influent COD/TN ratio should be less than 0.70. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Monoclonal antibody "gold rush".

    Science.gov (United States)

    Maggon, Krishan

    2007-01-01

    The market, sales and regulatory approval of new human medicines, during the past few years, indicates increasing number and share of new biologics and emergence of new multibillion dollar molecules. The global sale of monoclonal antibodies in 2006 were $20.6 billion. Remicade had annual sales gain of $1 billion during the past 3 years and five brands had similar increase in 2006. Rituxan with 2006 sales of $4.7 billion was the best selling monoclonal antibody and biological product and the 6th among the top selling medicinal brand. It may be the first biologic and monoclonal antibody to reach $10 billion annual sales in the near future. The strong demand from cancer and arthritis patients has surpassed almost all commercial market research reports and sales forecast. Seven monoclonal antibody brands in 2006 had sales exceeding $1 billion. Humanized or fully human monoclonal antibodies with low immunogenicity, enhanced antigen binding and reduced cellular toxicity provide better clinical efficacy. The higher technical and clinical success rate, overcoming of technical hurdles in large scale manufacturing, low cost of market entry and IND filing, use of fully human and humanized monoclonal antibodies has attracted funds and resources towards R&D. Review of industry research pipeline and sales data during the past 3 years indicate a real paradigm shift in industrial R&D from pharmaceutical to biologics and monoclonal antibodies. The antibody bandwagon has been joined by 200 companies with hundreds of new projects and targets and has attracted billions of dollars in R&D investment, acquisitions and licensing deals leading to the current Monoclonal Antibody Gold Rush.

  6. Serum herpes simplex antibodies

    Science.gov (United States)

    ... causes cold sores (oral herpes). HSV-2 causes genital herpes. How the Test is Performed A blood sample ... person has ever been infected with oral or genital herpes . It looks for antibodies to herpes simplex virus ...

  7. Anti-sulfotyrosine antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Bertozzi, Carolyn R [Berkeley, CA; Kehoe, John [Saint Davids, PA; Bradbury, Andrew M [Santa Fe, NM

    2009-09-15

    The invention provides anti-sulfotyrosine specific antibodies capable of detecting and isolating polypeptides that are tyrosine-sulfated. The sulfotyrosine antibodies and antibody fragments of the invention may be used to discriminate between the non-sulfated and sulfated forms of such proteins, using any number of immunological assays, such ELISAs, immunoblots, Western Blots, immunoprecipitations, and the like. Using a phage-display system, single chain antibodies (scFvs) were generated and screened against tyrosine-sulfated synthetic peptide antigens, resulting in the isolation of scFvs that specifically recognize sulfotyrosine-containing peptides and/or demonstrate sulfotyrosine-specific binding in tyrosine sulfated proteins. The VH and VL genes from one such sulfotyrosine-specific scFv were employed to generate a full length, sulfotyrosine-specific immunoglobulin.

  8. Bifunctional antibodies for radioimmunotherapy.

    Science.gov (United States)

    Chatal, J F; Faivre-Chauvet, A; Bardies, M; Peltier, P; Gautherot, E; Barbet, J

    1995-04-01

    In two-step targeting technique using bifunctional antibodies, a nonradiolabeled immunoconjugate with slow uptake kinetics (several days) is initially injected, followed by a small radiolabeled hapten with fast kinetics (several hours) that binds to the bispecific immunoconjugate already taken up by the tumor target. In patients with colorectal or medullary thyroid cancer, clinical studies performed with an anti-CEA/anti-DTPA-indium bifunctional antibody and an indium-111-labeled di-DTPA-TL bivalent hapten showed that tumor uptake was not modified compared to results for F(ab')2 fragments of the same anti-CEA antibody directly labeled with indium-111, whereas the radioactivity of normal tissues was significantly reduced (3- to 6-fold). The fast tumor uptake kinetics (several hours) and high or very high tumor-to-normal tissue ratios obtained with the bifunctional antibody technique are favorable parameters for efficient radioimmunotherapy.

  9. Antibody Blood Tests

    Science.gov (United States)

    Antibody Blood Tests Researchers have discovered that people with celiac disease who eat gluten have higher than normal levels of ... do I do if I have a negative blood test (or panel) but I’m still having symptoms? ...

  10. Glycan elongation beyond the mucin associated Tn antigen protects tumor cells from immune-mediated killing

    DEFF Research Database (Denmark)

    Madsen, Caroline B; Lavrsen, Kirstine; Steentoft, Catharina

    2013-01-01

    and pancreatic cancer cell lines T47D and Capan-1 increases sensitivity to both NK cell mediated antibody-dependent cellular-cytotoxicity (ADCC) and cytotoxic T lymphocyte (CTL)-mediated killing. In addition, we investigated the association between total cell surface expression of MUC1/MUC16 and NK or CTL...

  11. In vivo study of the HC-TN strain of hepatitis C virus recovered from a patient with fulminant hepatitis: RNA transcripts of a molecular clone (pHC-TN) are infectious in chimpanzees but not in Huh7.5 cells

    DEFF Research Database (Denmark)

    Sakai, Akito; Takikawa, Shingo; Thimme, Robert

    2007-01-01

    disease severity, host immune response, viral evolution, and outcome. A second chimpanzee (CH1581) was infected from CH1422 plasma, and a third chimpanzee (CH1579) was infected from RNA transcripts of a consensus cDNA of HC-TN (pHC-TN). RNA transcripts of pHC-TN did not replicate in Huh7.5 cells, which......Both viral and host factors are thought to influence the pathogenesis of hepatitis C virus (HCV) infection. We studied strain HC-TN (genotype 1a), which caused fulminant hepatic failure in a patient and, subsequently, severe hepatitis in a chimpanzee (CH1422), to analyze the relationship between...

  12. HTR-TN a European network for the development of HTR technology

    International Nuclear Information System (INIS)

    Von Lensa, W.

    2001-01-01

    A network called High-temperature reactor technology network (HTR-TN) has been created at a European level to coordinate works and knowledge on the subject with a long-term perspective and to serve as a channel for international collaboration. An analysis confirmed that the obvious economic penalty of HTR due to its low density power could be compensated by the combination of recent advances that may completely change the positioning of HTR on the energy market: -) the modular concept allowed to get a reactor free from core melt risk without intervention of any active safety system, implying a drastic simplification of the design of the reactor and the safety systems as well as a standardisation and potential for shop fabrication in series; -) the development of gas turbines, the efficiency of which increased, in 10 years, from 35% till 50% and more, enabling to consider suppression of the secondary system; -) the ultra high burn-up potential of HTR fuel and the possibility for direct disposal of spent HTR fuel elements that may reduce cost of the fuel cycle and contribute to the reduction of civil and military plutonium stockpiles. (A.C.)

  13. Characterization of the pJB12 Plasmid from Pseudomonas aeruginosa Reveals Tn6352, a Novel Putative Transposon Associated with Mobilization of the blaVIM-2-Harboring In58 Integron

    OpenAIRE

    Botelho, João; Grosso, Filipa; Peixe, Luísa

    2017-01-01

    The blaVIM-2-carrying In58 integron has been linked to a chromosomal location in different bacterial species, including Pseudomonas aeruginosa. This work reports the first fully sequenced In58-harboring plasmid, which is significantly different from the two previously identified blaVIM-2-carrying plasmids in P. aeruginosa. blaVIM-2 might have been acquired by transposition of Tn6352, a novel transposon composed of the In58 and ISPa17 elements. The recognition of similar inverted repeat (IR) s...

  14. The Breast Cancer-Associated Glycoforms of MUC1, MUC1-Tn and sialyl-Tn, Are Expressed in COSMC Wild-Type Cells and Bind the C-Type Lectin MGL

    DEFF Research Database (Denmark)

    Beatson, Richard; Maurstad, Gjertrud; Picco, Gianfranco

    2015-01-01

    that MUC1 carrying both Tn or STn can bind to the C-type lectin MGL and using atomic force microscopy show that they bind to MGL with a similar deadadhesion force. Tumour associated STn is associated with poor prognosis and resistance to chemotherapy in breast carcinomas, inhibition of DC maturation, DC...... carbohydrate binding lectins. Two glyco-epitopes that are found expressed by many carcinomas are Tn (GalNAc-Ser/Thr) and STn (NeuAcα2,6GalNAc-Ser/Thr). These glycans can be carried on many mucin-type glycoproteins including MUC1. We show that the majority of breast cancers carry Tn within the same cell......Aberrant glycosylation occurs in the majority of human cancers and changes in mucin-type O-glycosylation are key events that play a role in the induction of invasion and metastases. These changes generate novel cancer-specific glyco-antigens that can interact with cells of the immune system through...

  15. Sequence-Based Characterization of Tn5801-Like Genomic Islands in Tetracycline-Resistant Staphylococcus pseudintermedius and Other Gram-positive Bacteria from Humans and Animals

    DEFF Research Database (Denmark)

    de Vries, Lisbeth Elvira; Hasman, Henrik; Jurado Rabadán, Sonia

    2016-01-01

    Antibiotic resistance in pathogens is often associated with mobile genetic elements, such as genomic islands (GI) including integrative and conjugative elements (ICEs). These can transfer resistance genes within and between bacteria from humans and/or animals. The aim of this study was to investi......Antibiotic resistance in pathogens is often associated with mobile genetic elements, such as genomic islands (GI) including integrative and conjugative elements (ICEs). These can transfer resistance genes within and between bacteria from humans and/or animals. The aim of this study...... was to investigate whether Tn5801-like GIs carrying the tetracycline resistance gene, tet(M), are common in Staphylococcus pseudintermedius from pets, and to do an overall sequences-based characterization of Tn5801-like GIs detected in Gram-positive bacteria from humans and animals. A total of 27 tetracycline......(M). Out of 13 selected isolates, seven contained Tn5801-like GIs and six contained Tn916-like ICEs. Two different Tn5801-like GI types were detected among S. pseudintermedius (Tn5801 and GI6287) - both showed high similarity compared to GenBank sequences from human pathogens. Two distinct Tn5801-like GI...

  16. Natural and Man-made Antibody Repertories for Antibody Discovery

    Directory of Open Access Journals (Sweden)

    Juan C eAlmagro

    2012-11-01

    Full Text Available Antibodies are the fastest-growing segment of the biologics market. The success of antibody-based drugs resides in their exquisite specificity, high potency, stability, solubility, safety and relatively inexpensive manufacturing process in comparison with other biologics. We outline here the structural studies and fundamental principles that define how antibodies interact with diverse targets. We also describe the antibody repertoires and affinity maturation mechanisms of human, mice and chickens, plus the use of novel single-domain antibodies in camelids and sharks. These species all utilize diverse evolutionary solutions to generate specific and high affinity antibodies and illustrate the plasticity of natural antibody repertoires. In addition, we discuss the multiple variations of man-made antibody repertoires designed and validated in the last two decades, which have served as tools to explore how the size, diversity and composition of a repertoire impact the antibody discovery process.

  17. Molecular basis of antibody binding to mucin glycopeptides in lung cancer

    Science.gov (United States)

    QU, JIN; YU, HONGTAO; LI, FENGE; ZHANG, CHUNLEI; TRAD, AHMAD; BROOKS, CORY; ZHANG, BIN; GONG, TING; GUO, ZHI; LI, YUNSEN; RAGUPATHI, GOVIND; LOU, YANYAN; HWU, PATRICK; HUANG, WEI; ZHOU, DAPENG

    2016-01-01

    Glycopeptides bearing Tn epitopes are emerging targets for cancer diagnosis and immunotherapy. In this study, we analyzed membrane proteins containing O-glycosylated tandem repeat (TR) sequences in lung cancer patients of different types and stages, using gene microarray data in public domain. The expression of Tn and glycopeptide epitopes on the surface of lung cancer cell lines were studied by monoclonal IgG antibodies 14A, 16A, and B72.3. The binding of mAbs to synthetic glycopeptides were studied by surface plasmon resonance. Nine mucin mRNAs were found to be expressed in lung cancer patients but at similar level to healthy individuals. At protein level, a glycopeptide epitope on cancer cell surface is preferably recognized by mAb 16A, as compared to peptide-alone (14A) or sugar-alone epitopes (B72.3). 14A and 16A favor clustered TR containing more than three TR sequences, with 10-fold lower Kd than two consecutive TR. B72.3 preferrably recognized clustered sialyl-Tn displayed on MUC1 but not other O-glycoproteins, with 100-fold stronger binding when MUC1 is transfected as a sugar carrier, while the total sugar epitopes remain unchanged. These findings indicate that clusters of both TR backbones and sugars are essential for mAb binding to mucin glycopeptides. Three rules of antibody binding to mucin glycopeptides at molecular level are presented here: first, the peptide backbone of a glycopeptide is preferentially recognized by B cells through mutations in complementarity determining regions (CDRs) of B cell receptor, and the sugar-binding specificity is acquired through mutations in frame work of heavy chain; secondly, consecutive tandem repeats (TR) of peptides and glycopeptides are preferentially recognized by B cells, which favor clustered TR containing more than three TR sequences; thirdly, certain sugar-specific B cells recognize and accommodate clustered Tn and sialyl-Tn displayed on the surface of a mucin but not other membrane proteins. PMID:26692014

  18. First report of an OXA-23 carbapenemase-producing Acinetobacter baumannii clinical isolate related to Tn2006 in Spain.

    Science.gov (United States)

    Espinal, P; Macià, M D; Roca, I; Gato, E; Ruíz, E; Fernández-Cuenca, F; Oliver, A; Rodríguez-Baño, J; Bou, G; Tomás, M; Vila, J

    2013-01-01

    A carbapenem-resistant Acinetobacter baumannii clinical isolate belonging to European clone II and sequence type 2 was recovered from a patient in the Son Espases hospital in Mallorca, Spain. Genetic analysis showed the presence of the bla(OXA-23) gene in association with the widely disseminated transposon Tn2006. This is the first reported identification of A. baumannii carrying bla(OXA-23) in Spain.

  19. Polyclonal Antibody Production for Membrane Proteins via Genetic Immunization.

    Science.gov (United States)

    Hansen, Debra T; Robida, Mark D; Craciunescu, Felicia M; Loskutov, Andrey V; Dörner, Katerina; Rodenberry, John-Charles; Wang, Xiao; Olson, Tien L; Patel, Hetal; Fromme, Petra; Sykes, Kathryn F

    2016-02-24

    Antibodies are essential for structural determinations and functional studies of membrane proteins, but antibody generation is limited by the availability of properly-folded and purified antigen. We describe the first application of genetic immunization to a structurally diverse set of membrane proteins to show that immunization of mice with DNA alone produced antibodies against 71% (n = 17) of the bacterial and viral targets. Antibody production correlated with prior reports of target immunogenicity in host organisms, underscoring the efficiency of this DNA-gold micronanoplex approach. To generate each antigen for antibody characterization, we also developed a simple in vitro membrane protein expression and capture method. Antibody specificity was demonstrated upon identifying, for the first time, membrane-directed heterologous expression of the native sequences of the FopA and FTT1525 virulence determinants from the select agent Francisella tularensis SCHU S4. These approaches will accelerate future structural and functional investigations of therapeutically-relevant membrane proteins.

  20. Antibodies against HLA-DP recognize broadly expressed epitopes.

    Science.gov (United States)

    Simmons, Daimon P; Kafetzi, Maria L; Wood, Isabelle; Macaskill, Peter C; Milford, Edgar L; Guleria, Indira

    2016-12-01

    HLA matching and avoidance of pre-transplant donor-specific antibodies are important in selection of donors for solid organ transplant. Solid phase testing with single antigen beads allows resolution of antibody reactivity to the level of the allele. Single antigen bead testing results at a large transplant center were reviewed to identify selective reactivity patterns of anti-HLA antibodies. Many HLA-DP antibodies were identified in the context of other HLA antibodies, but some sera had antibodies against only HLA-DP. B cell flow crossmatch testing was positive for 2 out of 9 sera with HLA-DP antibodies. Many patterns of reactivity corresponded to epitopes in hypervariable regions C and F of DPB1, but some matched epitopes in other regions or DPA1. Through analysis of single antigen bead testing from a large number of patients, we report that anti-HLA-DP antibodies predominantly recognize broadly cross-reactive epitopes. The United Network for Organ Sharing has mandated HLA-DP typing on all deceased kidney donors, and HLA-DP epitopes should be considered as the major antigens for avoidance of pre-transplant donor-specific antibodies. Published by Elsevier Inc.

  1. Identification of antibody-interacting proteins that contribute to the production of recombinant antibody in mammalian cells.

    Science.gov (United States)

    Nishimiya, Daisuke; Ogura, Yuji; Sakurai, Hidetaka; Takahashi, Tohru

    2012-11-01

    Protein folding and assembly processes are essential for antibody secretion; however, the endogenous proteins involved in these processes remain largely unknown. Therefore, except for some well-known endoplasmic reticulum (ER) chaperones such as GRP78/Bip and protein disulfide isomerase, enhancement of recombinant antibody expression by co-expression of interacting proteins has been largely elusive. Here, in addition to known ER chaperones, we identified additional endogenous proteins that interact with recombinant antibody in mammalian cells by immunoprecipitation coupled with liquid chromatography-tandem mass spectrometry. Most of our identified proteins enhanced antibody production, and furthermore, some of their combinations resulted in greater enhancement. In particular, eukaryotic initiation factor 4A combined with other proteins had approximately fourfold higher effect on antibody production. Identified proteins that could improve antibody expression contain not only ER-resident proteins like GRP78/Bip but also non-ER-resident proteins. These results suggest that this method could be effective in the investigation of novel proteins that are involved in enhancing recombinant antibody production because immunoprecipitation coupled with mass spectroscopy could identify proteins which directly interact with the antibody.

  2. Antithyroglobulin Antibodies and Antimicrosomal Antibodies in Various Thyroid Diseases

    International Nuclear Information System (INIS)

    Lee, Gwon Jun; Hong, Key Sak; Choi, Kang Won; Lee, Kyu; Koh, Chang Soon; Lee, Mun Ho; Park, Sung Hoe; Chi, Je Geun; Lee, Sang Kook

    1979-01-01

    The authors investigated the incidence of antithyroglobulin antibodies and antibodies and antimicrosomal antibodies measured by tanned red cell hemagglutination method in subjects suffering from various thyroid disorders. 1) In 15 normal patients, neither suffering from any thyroid diseases nor from any other autoimmune disorders, the antithyroglobulin antibodies were all negative, but the antimicrosomal antibody was positive only in one patient (6.7%). 2) The antithyroglobulin antibodies were positive in 31.5% (34 patients) of 108 patients with various thyroid diseases, and the antimicrosomal antibodies were positive in 37.0% (40 patients). 3) of the 25 patients with Graves' diseases, 7 patients (28.0%) showed positive for the antithyroglobulin antibodies, and 9 (36.0%) for the antimicrosomal antibodies. There was no definite differences in clinical and thyroid functions between the groups with positive and negative results. 4) Both antibodies were positive in 16 (88.9%) and 17 (94.4%) patients respectively among 18 patients with Hashimoto's thyroiditis, all of them were diagnosed histologically. 5) Three out of 33 patients with thyroid adenoma showed positive antibodies, and 3 of 16 patients with thyroid carcinoma revealed positive antibodies. 6) TRCH antibodies demonstrated negative results in 2 patients with subacute thyroiditis, but positive in one patient with idiopathic primary myxedema. 7) The number of patients with high titers(>l:802) was 16 for antithyroglobulin antibody, and 62.5% (10 patients) of which was Hashimoto's thyroiditis. Thirteen (65.0) of 20 patients with high titers (>l:802) for antimicrosomal antibody was Hashimoto's thyroiditis. TRCH test is a simple, sensitive method, and has high reliability and reproducibility. The incidences and titers of antithyroglobulin antibody and antimicrosomal antibody are especially high in Hashimoto's thyroiditis.

  3. Human anti-animal antibody interferences in immunological assays.

    Science.gov (United States)

    Kricka, L J

    1999-07-01

    The scope and significance of human anti-animal antibody interference in immunological assays is reviewed with an emphasis on human anti-animal immunoglobulins, particularly human anti-mouse antibodies (HAMAs). Anti-animal antibodies (IgG, IgA, IgM, IgE class, anti-isotype, and anti-idiotype specificity) arise as a result of iatrogenic and noniatrogenic causes and include human anti-mouse, -rabbit, -goat, -sheep, -cow, -pig, -rat, and -horse antibodies and antibodies with mixed specificity. Circulating antibodies can reach gram per liter concentrations and may persist for years. Prevalence estimates for anti-animal antibodies in the general population vary widely and range from HAMA, which causes both positive and negative interferences in two-site mouse monoclonal antibody-based assays. Strategies to prevent the development of human anti-animal antibody responses include immunosuppressant therapy and the use of humanized, polyethylene glycolylated, or Fab fragments of antibody agents. Sample pretreatment or assay redesign can eliminate immunoassay interferences caused by anti-animal antibodies. Enzyme immunoassays, immunoradiometric assays, immunofluorescence, and HPLC assays have been designed to detect HAMA and other anti-animal antibodies, but intermethod comparability is complicated by differences in assay specificity and lack of standardization. Human anti-animal antibodies often go unnoticed, to the detriment of patient care. A heightened awareness on the part of laboratory staff and clinicians of the problems caused by this type of interference in routine immunoassay tests is desirable. Efforts should be directed at improving methods for identifying and eliminating this type of analytical interference.

  4. Compositions, antibodies, asthma diagnosis methods, and methods for preparing antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Hongjun; Zangar, Richard C.

    2017-01-17

    Methods for preparing an antibody are provided with the method including incorporating 3-bromo-4-hydroxy-benzoic acid into a protein to form an antigen, immunizing a mammalian host with the antigen, and recovering an antibody having an affinity for the antigen from the host. Antibodies having a binding affinity for a monohalotyrosine are provided as well as composition comprising an antibody bound with monohalotyrosine. Compositions comprising a protein having a 3-bromo-4-hydroxy-benzoic acid moiety are also provided. Methods for evaluating the severity of asthma are provide with the methods including analyzing sputum of a patient using an antibody having a binding affinity for monohalotyrosine, and measuring the amount of antibody bound to protein. Methods for determining eosinophil activity in bodily fluid are also provided with the methods including exposing bodily fluid to an antibody having a binding affinity for monohalotyrosine, and measuring the amount of bound antibody to determine the eosinophil activity.

  5. Monoclonal antibodies technology. Protocols

    International Nuclear Information System (INIS)

    Acevado Castro, B.E.

    1997-01-01

    reached at least 3/4 confluency, 2 ampules/cell culture. Save the supernatants for further studies. Regarding to separate the interested cells of irrelevant cells, you must been do a limited diluted procedures. Ascitis production. To produce ascitis the number of cells depend of the every particular clone. Usually we inoculated ip between 1 to 2 x10 6 cells/ml/mice in PBS buffer. The animals must be inoculated intraperitonealy with Pristan (Sigma) or mineral oil using 0.5 ml per mice. The inoculation could be occurred 7 to 10 days before to inoculated the clone. The weight of the animals must be controlled in order to obtain better yield. The bioreactors are an alternative method to produce AcMs without it have not used animals models. IgG purification process on protein A columns. Mouse antibodies of the IgG 1 subclass do not have a high affinity for protein A. Every subclass could be eluted at different peaks using a gradient pH. Usually we use 3M NaCl/1.5 M Gly pH 8.9 as a coupling buffer. For eluting IgGs we use 0.1M Citric Acid in a gradient pH (6 to IgG 1 , 5 to IgG 2a or IgG 2b , 4 IgG 3 ). 1. Adjust a Sepharose Protein A (Pharmacia) beads using NaCl/1.5 M Gly pH 8.9 (coupling buffer). 2. Diluted the ascitis 1:10 in coupling buffer using a filter paper for remove the aggregations. 3. Pass the solution trough a Protein A bead column at a standarized flow according your equipment. The capacity of the Protein A beads for IgGl is approximately 5 mg/ml of wet beads. Ascitis contain between 1-10 mg/ml. 4. Wash the colums with 10 volumes of coupling buffer. 5. Elute the column with 0.1 M Citric Acid in a gradient pH starting at 6 to 4. Collect the eluate in appropriate tube, and identify the immunoglobulin-containing fractions by absorbance a 280 nm (1 DO = 0.8 mg/ml)

  6. Chromosomal insertion of the entire Escherichia coli lactose operon, into two strains of Pseudomonas, using a modified mini-Tn5 delivery system

    DEFF Research Database (Denmark)

    Hansen, L. H.; Sørensen, S. J.; Jensen, Lars Bogø

    1997-01-01

    A 12-kb PstI fragment including the entire E. coli lactose operon (lacIPOZYA) was inserted in one copy into the chromosome of Pseudomonas putida, Pseudomonas fluorescens and an E. coli strain with lac(-) phenotype. This was made possible by improvements of an already existing mini-Tn5 transposon...... flanked by NotI sites needed in the mini-Tn5 delivery system; (b) the generation of E. coli nonlysogenic strains expressing the pi protein thus being capable of maintaining and delivering R6K-based mini-Tn5 vectors to other E. coli strains; (c) the successful insertion of the E. coli lactose operon...... into the P. fluorescens chromosome giving P. fluorescens the ability to grow on lactose; (d) evidence from Southern blotting that contradicts the assumption that the mini-Tn5 delivery system always creates one-copy inserts. These improvements allow insertion of large DNA fragments encoding highly expressed...

  7. Clinical relevance of determination the changes of serum cTnI, NPY and PGMP contents after treatment in patients with coronary heart diseases

    International Nuclear Information System (INIS)

    Ge Hengsong; Ma Chao; Wu Kaixia

    2011-01-01

    Objective: To explore the clinical significance of changes of serum cTnI, NPY and PGMP levels in patients with coronary heart diseases after treatment. Methods: Serum NPY, PGMP (with RIA), serum cTnI (with Biochemistry) levels were measured in 41 patients with coronary heart diseases both before and after treatment as well as in 35 normal controls. Results: Before treatment, serum cTnI, NPY and PGMP levels were significantly higher than those in controls (P 0.05), but serum PGMP level remained prominently higher (P<0.05). Conclusion: Pathogenesis and progress of CHD, might be closely related to serum cTnI, NPY and PGMP levels. (authors)

  8. Antithyroid microsomal antibody

    Science.gov (United States)

    ... that you have a higher chance of developing thyroid disease in the future. Antithyroid microsomal antibodies may be ... PA: Elsevier; 2016:chap 11. Weiss RE, Refetoff S. Thyroid function testing. In: Jameson JL, De Groot LJ, eds. Endocrinology: Adult and ... Lupus Read more ...

  9. Antibodies Targeting EMT

    Science.gov (United States)

    2017-10-01

    determine their targets on the cell. The newly discovered antibodies will then be engineered for utility as new highly specific drugs and diagnostics in...are from the aldo-keto reductase family (AKRs). Remarkably, 3 of the top 10 genes with induction in the mesenchymal TES2b cells Figure 1. Amino

  10. Monoclonal antibodies in haematopathology

    Energy Technology Data Exchange (ETDEWEB)

    Grignani, F.; Martelli, M.F.; Mason, D.Y.

    1985-01-01

    This book contains over 40 selections. Some of the titles are: Oncogene (c-myc, c-myb) amplification in acute myelogenous leukaemia; Ultrastructural characterization of leukaemic cells with monoloclonal antibodies; Origin of B-cell malignancies; Immunohistology of gut lymphomas; and Spurious evidence of lineage infidelity in monocytic leukaemia.

  11. Monoclonal antibodies in myeloma

    DEFF Research Database (Denmark)

    Sondergeld, P.; van de Donk, N. W. C. J.; Richardson, P. G.

    2015-01-01

    The development of monoclonal antibodies (mAbs) for the treatment of disease goes back to the vision of Paul Ehrlich in the late 19th century; however, the first successful treatment with a mAb was not until 1982, in a lymphoma patient. In multiple myeloma, mAbs are a very recent and exciting add...

  12. Sex-specific 99th percentiles derived from the AACC Universal Sample Bank for the Roche Gen 5 cTnT assay: Comorbidities and statistical methods influence derivation of reference limits.

    Science.gov (United States)

    Gunsolus, Ian L; Jaffe, Allan S; Sexter, Anne; Schulz, Karen; Ler, Ranka; Lindgren, Brittany; Saenger, Amy K; Love, Sara A; Apple, Fred S

    2017-12-01

    Our purpose was to determine a) overall and sex-specific 99th percentile upper reference limits (URL) and b) influences of statistical methods and comorbidities on the URLs. Heparin plasma from 838 normal subjects (423 men, 415 women) were obtained from the AACC (Universal Sample Bank). The cobas e602 measured cTnT (Roche Gen 5 assay); limit of detection (LoD), 3ng/L. Hemoglobin A1c (URL 6.5%), NT-proBNP (URL 125ng/L) and eGFR (60mL/min/1.73m 2 ) were measured, along with identification of statin use, to better define normality. 99th percentile URLs were determined by the non-parametric (NP), Harrell-Davis Estimator (HDE) and Robust (R) methods. 355 men and 339 women remained after exclusions. Overallstatistical method used influenced URLs as follows: pre/post exclusion overall, NP 16/16ng/L, HDE 17/17ng/L, R not available; men NP 18/16ng/L, HDE 21/19ng/L, R 16/11ng/L; women NP 13/10ng/L, HDE 14/14ng/L, R not available. We demonstrated that a) the Gen 5 cTnT assay does not meet the IFCC guideline for high-sensitivity assays, b) surrogate biomarkers significantly lowers the URLs and c) statistical methods used impact URLs. Our data suggest lower sex-specific cTnT 99th percentiles than reported in the FDA approved package insert. We emphasize the importance of detailing the criteria used to include and exclude subjects for defining a healthy population and the statistical method used to calculate 99th percentiles and identify outliers. Copyright © 2017 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  13. Intravenous avidin chase improved localization of radiolabeled streptavidin in intraperitoneal xenograft pretargeted with biotinylated antibody

    International Nuclear Information System (INIS)

    Zhang Meili; Sakahara, Harumi; Yao Zhengsheng; Saga, Tsuneo; Nakamoto, Yuhi; Sato, Noriko; Nakada, Hiroshi; Yamashina, Ikuo; Konishi, Junji

    1997-01-01

    In the present study, we examined the effect of avidin administered intravenously (i.v.) on the biodistribution of radiolabeled streptavidin in mice bearing intraperitoneal (IP) xenografts pretargeted with biotinylated antibody. Tumors were established in nude mice by IP inoculation of LS180 human colon cancer cells. Monoclonal antibody MLS128, which recognizes Tn antigen on mucin, was biotinylated and injected IP into the IP tumor-bearing mice. Radioiodinated streptavidin was administered IP or i.v. 48 h after pretargeting of biotinylated antibody. Avidin was administered i.v. 30 min prior to streptavidin injection. The localization of radioiodinated streptavidin in the tumor pretargeted with biotinylated antibody was significantly higher than that without pretargeting and that of radioiodinated MLS128 by the one-step method. Avidin administration significantly accelerated the clearance of radioiodinated streptavidin in blood and other normal tissues and increased the tumor-to-blood radioactivity ratio regardless of administration route of streptavidin. The i.v. avidin chase improved tumor localization of radiolabeled streptavidin in the IP xenografts pretargeted with biotinylated antibody

  14. Identifying sarcopenia.

    Science.gov (United States)

    Abellan van Kan, Gabor; Houles, Mathieu; Vellas, Bruno

    2012-09-01

    The present review describes and discusses the currently available definitions for sarcopenia from consensus studies. Different sarcopenia definitions have been proposed in these last years. Six main approaches to an operative definition of sarcopenia have been identified. Although the first definitions were solely based on the assessment of the amount of muscle mass, current definitions seem to consistently recognize a bi-dimensional nature of sarcopenia. So, these approaches imply the need of simultaneously assessing both age-related quantitative (i.e. amount of muscle mass) and qualitative (i.e. muscle strength and function) declines of skeletal muscle. Although current consensus exists about a bi-dimensional nature, the proposed approaches to measure sarcopenia are characterized by methodological differences. The majority of the operative definitions proposes to assess muscle mass as an index of appendicular muscle mass divided by squared height (evaluated by dual energy X-ray absorptiometry), assess strength using hand-held dynamometers, and assess function by evaluating gait speed at habitual pace over a short distance. Nevertheless, the clinically relevant thresholds and how to combine the three aspects in an operative definition in order to identify sarcopenia are heterogeneous. A main drawback is that supportive empirical data are missing for these conceptual definitions regarding the risk-assessment of different clinically significant adverse outcomes.

  15. Humanized Antibodies for Antiviral Therapy

    Science.gov (United States)

    Co, Man Sung; Deschamps, Marguerite; Whitley, Richard J.; Queen, Cary

    1991-04-01

    Antibody therapy holds great promise for the treatment of cancer, autoimmune disorders, and viral infections. Murine monoclonal antibodies are relatively easy to produce but are severely restricted for therapeutic use by their immunogenicity in humans. Production of human monoclonal antibodies has been problematic. Humanized antibodies can be generated by introducing the six hypervariable regions from the heavy and light chains of a murine antibody into a human framework sequence and combining it with human constant regions. We humanized, with the aid of computer modeling, two murine monoclonal antibodies against herpes simplex virus gB and gD glycoproteins. The binding, virus neutralization, and cell protection results all indicate that both humanized antibodies have retained the binding activities and the biological properties of the murine monoclonal antibodies.

  16. First results from TN273 studies of the SE Mariana Forearc rift

    Science.gov (United States)

    Ribeiro, J. M.; Stern, R. J.; Kelley, K. A.; Shaw, A. M.; Shimizu, N.; Martinez, F.; Ishii, T.; Ishizuka, O.; Manton, W. I.

    2012-12-01

    TN 273 aboard R/V Thomas Thompson (Dec. 22 2011- Jan. 22 2012) studied an unusual region of rifting affecting the southern Mariana forearc S.W. of Guam. The S.E. Mariana Forearc Rift (SEMFR) formed by diffuse tectonic and volcanic deformation (Martinez and Sleeper, this meeting) ~2.7-3.7 Ma ago to accommodate opening of the southernmost Mariana Trough backarc basin. A total of 730 km linear-track of SEMFR seafloor was surveyed with deep-towed side-scan sonar IMI-30. 14 dredges provided samples of SEMFR igneous rocks, analyzed for whole rock (WR) and glass compositions. These new results coupled with results of earlier investigations confirm that SEMFR is dominated by Miocene lavas along with minor gabbro and diabase. SEMFR lavas range in major element composition from primitive basalt to fractionated andesite (Mg# = 0.36-0.73; SiO2 = 50-57 wt%), mainly controlled by crystal fractionation. Rare Earth Element (REE) patterns range from LREE-depleted, N-MORB-like to flat patterns, reflecting different mantle processes (i.e. different sources, degree of melting …). Glassy rinds and olivine-hosted melt inclusions in these lavas contain variable volatile compositions (F = 75-358 ppm, S = 35-1126 ppm, Cl= 74-1400 ppm, CO2 = 15-520 ppm, 0.36-2.36 wt% H2O). SEMFR lavas show spider diagrams with positive anomalies in LILE and negative anomalies in HSFE. SEMFR lavas have backarc basin-like (BAB-like) chemical composition (H2O < 2.5wt%, Ba/Yb~20, Nb/Yb~1 and ɛNd~9) along with stronger enrichment in Rb and Cs than arc and BAB lavas, as demonstrated by their higher Rb/Th and Cs/Ba ratios in WR and glasses, which may reflect the role of the ultra-shallow fluids. Ultra-shallow fluids are derived from the top of the subducting slab, beneath the forearc, where most of the water and the fluid-mobile elements (Rb, Cs, Ba,) are thought to be released (Schmidt and Poli, 1998, EPSL, Savov et al., 2005, G-3). Our results suggest that i) SEMFR lavas formed by metasomatism of a BAB mantle

  17. Construction of phage antibody repertoires from the blood of West Nile virus-infected donors.

    Science.gov (United States)

    Throsby, Mark; de Kruif, John

    2009-01-01

    A method for the construction of West Nile virus immune donor antibody repertoires is described. B cells are harvested from a suitable donor and the antibody variable genes are amplified using polymerase chain reaction (PCR). The PCR fragments are cloned in a phage display vector to construct a repertoire that can be used in panning procedures to identify many unique monoclonal antibodies.

  18. Potential for novel MUC1 glycopeptide-specific antibody in passive cancer immunotherapy

    DEFF Research Database (Denmark)

    Madsen, Caroline B; Wandall, Hans H; Pedersen, Anders Elm

    2013-01-01

    MUC1 is an important target for antibodies in passive cancer immunotherapy. Antibodies against mucin glycans or mucin peptide backbone alone may give rise to cross reactivity with normal tissues. Therefore, attempts to identify antibodies against cancer-specific MUC1 glycopeptide epitopes havebeen...

  19. Optimization of high-rate TN removal in a novel constructed wetland integrated with microelectrolysis system treating high-strength digestate supernatant.

    Science.gov (United States)

    Guo, Luchen; He, Keli; Wu, Shubiao; Sun, Hao; Wang, Yanfei; Huang, Xu; Dong, Renjie

    2016-08-01

    The potential of high-rate TN removal in three aerated horizontal subsurface-flow constructed wetlands to treat high-strength anaerobic digestate supernatant was evaluated. Different strategies of intermittent aeration and effluent recirculation were applied to compare their effect on nitrogen depuration performance. Additional glucose supply and iron-activated carbon based post-treatment systems were established and examined, respectively, to further remove nitrate that accumulated in the effluents from aerated wetlands. The results showed that intermittent aeration (1 h on:1 h off) significantly improved nitrification with ammonium removal efficiency of 90% (18.1 g/(m(2)·d)), but limited TN removal efficiency (53%). Even though effluent recirculation (a ratio of 1:1) increased TN removal from 53% to 71%, the effluent nitrate concentration was still high. Additional glucose was used as a post-treatment option and further increased the TN removal to 82%; however, this implementation caused additional organic pollution. Furthermore, the iron-activated carbon system stimulated with a microelectrolysis process achieved greater than 85% effluent nitrate removal and resulted in 86% TN removal. Considering the high TN removal rate, aerated constructed wetlands integrated with a microelectrolysis-driven system show great potential for treating high-strength digestate supernatant. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Rapid and efficient introduction of a foreign gene into bacterial artificial chromosome-cloned varicella vaccine by Tn7-mediated site-specific transposition

    International Nuclear Information System (INIS)

    Somboonthum, Pranee; Koshizuka, Tetsuo; Okamoto, Shigefumi; Matsuura, Masaaki; Gomi, Yasuyuki; Takahashi, Michiaki; Yamanishi, Koichi; Mori, Yasuko

    2010-01-01

    Using a rapid and reliable system based on Tn7-mediated site-specific transposition, we have successfully constructed a recombinant Oka varicella vaccine (vOka) expressing the mumps virus (MuV) fusion protein (F). The backbone of the vector was our previously reported vOka-BAC (bacterial artificial chromosome) genome. We inserted the transposon Tn7 attachment sequence, LacZα-mini-attTn7, into the region between ORF12 and ORF13 to generate a vOka-BAC-Tn genome. The MuV-F expressing cassette was transposed into the vOka-BAC genome at the mini-attTn7 transposition site. MuV-F protein was expressed in recombinant virus, rvOka-F infected cells. In addition, the MuV-F protein was cleaved in the rvOka-F infected cells as in MuV-infected cells. The growth of rvOka-F was similar to that of the original recombinant vOka without the F gene. Thus, we show that Tn7-mediated transposition is an efficient method for introducing a foreign gene expression cassette into the vOka-BAC genome as a live virus vector.

  1. HCF-1 encoded by baculovirus AcMNPV is required for productive nucleopolyhedrovirus infection of non-permissive Tn368 cells.

    Science.gov (United States)

    Tachibana, Ami; Hamajima, Rina; Tomizaki, Moe; Kondo, Takuya; Nanba, Yoshie; Kobayashi, Michihiro; Yamada, Hayato; Ikeda, Motoko

    2017-06-19

    Baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) replicates in both Spodoptera frugiperda Sf21 and Trichoplusia ni Tn368 cells, whereas AcMNPV defective in hcf-1 (host cell-factor 1) gene productively infects only Sf21 cells, indicating that HCF-1 is indispensable for the AcMNPV productive infection of Tn368 cells. Here, we demonstrated that HCF-1 protein transiently expressed in Tn368 cells promotes the DNA synthesis of Hyphantria cunea MNPV (HycuMNPV), Orygia pseudotsugata MNPV and Bombyx mori NPV, which are normally unable to replicate in Tn368 cells. We also demonstrated that a recombinant HycuMNPV harboring the hcf-1 gene successfully replicates in Tn368 cells, generating substantial yields of progeny viruses and polyhedra. These results indicate that HCF-1 encoded by AcMNPV is an essential viral factor for productive NPV infection of Tn368 cells. Taken together with the previous findings on HRF-1 (host range factor 1), the present results provide strong evidence that viral genes acquired through horizontal gene transfer play an important role in baculovirus evolution, serving to expand the host range of baculoviruses.

  2. Multiple functional therapeutic effects of TnP: A small stable synthetic peptide derived from fish venom in a mouse model of multiple sclerosis.

    Science.gov (United States)

    Komegae, Evilin Naname; Souza, Tais Aparecida Matozo; Grund, Lidiane Zito; Lima, Carla; Lopes-Ferreira, Monica

    2017-01-01

    The pathological condition of multiple sclerosis (MS) relies on innate and adaptive immunity. New types of agents that beneficially modify the course of MS, stopping the progression and repairing the damage appear promising. Here, we studied TnP, a small stable synthetic peptide derived from fish venom in the control of inflammation and demyelination in experimental autoimmune encephalomyelitis as prophylactic treatment. TnP decreased the number of the perivascular infiltrates in spinal cord, and the activity of MMP-9 by F4/80+ macrophages were decreased after different regimen treatments. TnP reduces in the central nervous system the infiltration of IFN-γ-producing Th1 and IL-17A-producing Th17 cells. Also, treatment with therapeutic TnP promotes the emergence of functional Treg in the central nervous system entirely dependent on IL-10. Therapeutic TnP treatment accelerates the remyelination process in a cuprizone model of demyelination. These findings support the beneficial effects of TnP and provides a new therapeutic opportunity for the treatment of MS.

  3. Multiple functional therapeutic effects of TnP: A small stable synthetic peptide derived from fish venom in a mouse model of multiple sclerosis.

    Directory of Open Access Journals (Sweden)

    Evilin Naname Komegae

    Full Text Available The pathological condition of multiple sclerosis (MS relies on innate and adaptive immunity. New types of agents that beneficially modify the course of MS, stopping the progression and repairing the damage appear promising. Here, we studied TnP, a small stable synthetic peptide derived from fish venom in the control of inflammation and demyelination in experimental autoimmune encephalomyelitis as prophylactic treatment. TnP decreased the number of the perivascular infiltrates in spinal cord, and the activity of MMP-9 by F4/80+ macrophages were decreased after different regimen treatments. TnP reduces in the central nervous system the infiltration of IFN-γ-producing Th1 and IL-17A-producing Th17 cells. Also, treatment with therapeutic TnP promotes the emergence of functional Treg in the central nervous system entirely dependent on IL-10. Therapeutic TnP treatment accelerates the remyelination process in a cuprizone model of demyelination. These findings support the beneficial effects of TnP and provides a new therapeutic opportunity for the treatment of MS.

  4. Genome-wide investigation of the genes involved in nicotine metabolism in Pseudomonas putida J5 by Tn5 transposon mutagenesis.

    Science.gov (United States)

    Xia, Zhenyuan; Zhang, Wei; Lei, Liping; Liu, Xingzhong; Wei, Hai-Lei

    2015-08-01

    Pseudomonas putida J5 is an efficient nicotine-degrading bacterial strain isolated from the tobacco rhizosphere. We successfully performed a comprehensive whole-genome analysis of nicotine metabolism-associated genes by Tn5 transposon mutagenesis in P. putida J5. A total of 18 mutants with unique insertions screened from 16,324 Tn5-transformants failed to use nicotine as the sole carbon source. Flanking sequences of the Tn5 transposon were cloned with a shotgun method from all of the nicotine-growth-deficient mutants. The potentially essential products of mutated gene were classified as follows: oxidoreductases, protein and metal transport systems, proteases and peptidases, transcriptional and translational regulators, and unknown proteins. Bioinformatic analysis of the Tn5 insertion sites indicated that the nicotine metabolic genes were separated and widely distributed in the genome. One of the mutants, M2022, was a Tn5 insert into a gene encoding a homolog of 6-hydroxy-L-nicotine oxidase, the second enzyme of nicotine metabolism in Arthrobacter nicotinovorans. Genetic and biochemical analysis confirmed that three open reading frames (ORFs) from an approximately 13-kb fragment recovered from the mutant M2022 were responsible for the transformation of nicotine to 3-succinoyl-pyridine via pseudooxynicotine and 3-succinoyl semialdehyde-pyridine, the first three steps of nicotine degradation. Further research on these mutants and the Tn5-inserted genes will help us characterize nicotine metabolic processes in P. putida J5.

  5. Structural Characterization of Peptide Antibodies

    DEFF Research Database (Denmark)

    Chailyan, Anna; Marcatili, Paolo

    2015-01-01

    The role of proteins as very effective immunogens for the generation of antibodies is indisputable. Nevertheless, cases in which protein usage for antibody production is not feasible or convenient compelled the creation of a powerful alternative consisting of synthetic peptides. Synthetic peptides...... can be modified to obtain desired properties or conformation, tagged for purification, isotopically labeled for protein quantitation or conjugated to immunogens for antibody production. The antibodies that bind to these peptides represent an invaluable tool for biological research and discovery...

  6. Pre-existing Antibody: Biotherapeutic Modality-Based Review.

    Science.gov (United States)

    Gorovits, Boris; Clements-Egan, Adrienne; Birchler, Mary; Liang, Meina; Myler, Heather; Peng, Kun; Purushothama, Shobha; Rajadhyaksha, Manoj; Salazar-Fontana, Laura; Sung, Crystal; Xue, Li

    2016-03-01

    Pre-existing antibodies to biotherapeutic drugs have been detected in drug-naïve subjects for a variety of biotherapeutic modalities. Pre-existing antibodies are immunoglobulins that are either specific or cross-reacting with a protein or glycan epitopes on a biotherapeutic compound. Although the exact cause for pre-existing antibodies is often unknown, environmental exposures to non-human proteins, glycans, and structurally similar products are frequently proposed as factors. Clinical consequences of the pre-existing antibodies vary from an adverse effect on patient safety to no impact at all and remain highly dependent on the biotherapeutic drug modality and therapeutic indication. As such, pre-existing antibodies are viewed as an immunogenicity risk factor requiring a careful evaluation. Herein, the relationships between biotherapeutic modalities to the nature, prevalence, and clinical consequences of pre-existing antibodies are reviewed. Initial evidence for pre-existing antibody is often identified during anti-drug antibody (ADA) assay development. Other interfering factors known to cause false ADA positive signal, including circulating multimeric drug target, rheumatoid factors, and heterophilic antibodies, are discussed.

  7. The antibody horror show: an introductory guide for the perplexed.

    Science.gov (United States)

    Goodman, Simon L

    2018-02-02

    The biological literature reverberates with the inadequacies of commercial research-tool antibodies. The scientific community spends some $2 billion per year on such reagents. Excellent accessible scientific platforms exist for reliably making, validating and using antibodies, yet the laboratory end-user reality is somehow depressing - because they often "don't work". This experience is due to a bizarre and variegated spectrum of causes including: inadequately identified antibodies; inappropriate user and supplier validation; poor user training; and overloaded publishers. Colourful as this may appear, the outcomes for the community are uniformly grim, including badly damaged scientific careers, wasted public funding, and contaminated literature. As antibodies are amongst the most important of everyday reagents in cell biology and biochemistry, I have tried here to gently suggest a few possible solutions, including: a move towards using recombinant antibodies; obligatory unique identification of antibodies, their immunogens, and their producers; centralized international banking of standard antibodies and their ligands; routine, accessible open-source documentation of user experience with antibodies; and antibody-user certification. Copyright © 2018. Published by Elsevier B.V.

  8. Synthetic peptides for antibody production

    NARCIS (Netherlands)

    N.D. Zegers (Netty)

    1995-01-01

    textabstractSynthetic peptides are useful tools for the generation of antibodies. The use of antibodies as specific reagents in inununochemical assays is widely applied. In this chapter, the application of synthetic peptides for the generation of antibodies is described. The different steps

  9. Synthetic peptides for antibody production

    NARCIS (Netherlands)

    Zegers, N.D.

    1995-01-01

    Synthetic peptides are useful tools for the generation of antibodies. The use of antibodies as specific reagents in inununochemical assays is widely applied. In this chapter, the application of synthetic peptides for the generation of antibodies is described. The different steps that lead to the

  10. Magnetic Purification of Antibodies

    Science.gov (United States)

    Dhadge, Vijaykumar Laxman

    This work aimed at the development of magnetic nanoparticles for antibody purification and at the evaluation of their performance in Magnetic fishing and in a newly developed hybrid technology Magnetic Aqueous Two Phase Systems. Magnetic materials were produced by coprecipitation and solvothermal approaches. Natural polymers such as dextran, extracellular polysaccharide and gum Arabic were employed for coating of iron oxide magnetic supports. Polymer coated magnetic supports were then modified with synthetic antibody specific ligands,namely boronic acid, a triazine ligand (named 22/8) and an Ugi ligand (named A2C7I1). To optimize the efficacy of magnetic nanoparticles for antibody magnetic fishing, various solutions of pure and crude antibody solutions along with BSA as a non-specific binding protein were tested. The selectivity of magnetic nanoparticle for antibody, IgG, was found effective with boronic acid and ligand 22/8. Magnetic supports were then studied for their performance in high gradient magnetic separator for effective separation capability as well as higher volume handling capability. The magnetic materials were also supplemented to aqueous two phase systems, devising a new purification technology. For this purpose, magnetic particles modified with boronic acid were more effective. This alternative strategy reduced the time of operation,maximized separation capability (yield and purity), while reducing the amount of salt required. Boronic acid coated magnetic particles bound 170 +/- 10 mg hIgG/g MP and eluted 160 +/- 5 mg hIgG/g MP, while binding only 15 +/- 5 mg BSA/g MP. The affinity constant for the interaction between hIgG and APBA_MP was estimated as 4.9 x 105 M-1 (Ka) with a theoretical maximum capacity of 492 mg hIgG adsorbed/g MP (Qmax). APBA_MPs were also tested for antibody purification directly from CHO cell supernatants. The particles were able to bind 98% of IgG loaded and to recover 95% of pure IgG (purity greater than 98%) at extremely

  11. Human antibody and antigen response to IncA antibody of Chlamydia trachomatis.

    Science.gov (United States)

    Tsai, P Y; Hsu, M C; Huang, C T; Li, S Y

    2007-01-01

    The high prevalence of C. trachomatis worldwide has underscored the importance of identifying specific immunogenic antigens in facilitating diagnosis as well as vaccine development. The aim of this study is to evaluate IncA antibody and antigen production in natural human infections. Our temporal expression study showed that IncA transcription and protein expression could be detected as early as 4 hours after the start of infection. Antibody responses could be detected in urine and genital swab samples from C. trachomatis-positive patients. It is especially interesting to note that the IncA antigen could be detected in urine. In conclusion, we have identified IncA as an important antigen in human. The potential applicability of the IncA antibody or antigen in the diagnosis as well as to vaccine development for C. trachomatis is also discussed.

  12. Platelet antibody: review of detection methods

    Energy Technology Data Exchange (ETDEWEB)

    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.

  13. Clinical use of antibodies

    International Nuclear Information System (INIS)

    Baum, R.P.; Hoer, Gustav; Cox, P.H.; Buraggi, G.L.

    1991-01-01

    Use of monoclonal antibodies as tumour specific carrier molecules for therapeutic agents or as in vivo diagnostic reagents when labelled with radionuclides or NMR signal enhancers is attracting more and more attention. The potential is enormous but the technical problems are also considerable requiring the concerted action of many different scientific disciplines. This volume is based upon a symposium organised in Frankfurt in 1990 under the auspices of the European Association of Nuclear Medicines' Specialist Task Groups on Cardiology and the Utility of Labelled Antibodies. It gives a multidisciplinary review of the state of the art and of problems to be solved as well as recording the not inconsiderable successes which have been booked to date. The book will be of value as a reference to both clinicians and research scientists. refs.; figs.; tabs

  14. The Human Antibody Response to Dengue Virus Infection

    Directory of Open Access Journals (Sweden)

    Aravinda M. de Silva

    2011-11-01

    Full Text Available Dengue viruses (DENV are the causative agents of dengue fever (DF and dengue hemorrhagic fever (DHF. Here we review the current state of knowledge about the human antibody response to dengue and identify important knowledge gaps. A large body of work has demonstrated that antibodies can neutralize or enhance DENV infection. Investigators have mainly used mouse monoclonal antibodies (MAbs to study interactions between DENV and antibodies. These studies indicate that antibody neutralization of DENVs is a “multi-hit” phenomenon that requires the binding of multiple antibodies to neutralize a virion. The most potently neutralizing mouse MAbs bind to surface exposed epitopes on domain III of the dengue envelope (E protein. One challenge facing the dengue field now is to extend these studies with mouse MAbs to better understand the human antibody response. The human antibody response is complex as it involves a polyclonal response to primary and secondary infections with 4 different DENV serotypes. Here we review studies conducted with immune sera and MAbs isolated from people exposed to dengue infections. Most dengue-specific antibodies in human immune sera are weakly neutralizing and bind to multiple DENV serotypes. The human antibodies that potently and type specifically neutralize DENV represent a small fraction of the total DENV-specific antibody response. Moreover, these neutralizing antibodies appear to bind to novel epitopes including complex, quaternary epitopes that are only preserved on the intact virion. These studies establish that human and mouse antibodies recognize distinct epitopes on the dengue virion. The leading theory proposed to explain the increased risk of severe disease in secondary cases is antibody dependent enhancement (ADE, which postulates that weakly neutralizing antibodies from the first infection bind to the second serotype and enhance infection of FcγR bearing myeloid cells such as monocytes and macrophages. Here

  15. Effect of subinhibitory concentrations of four commonly used biocides on the conjugative transfer of Tn916 in Bacillus subtilis

    DEFF Research Database (Denmark)

    Seier-Petersen, Maria Amalie; Jasni, A.; Aarestrup, Frank Møller

    2014-01-01

    Objectives Large amounts of biocides are used to reduce and control bacterial growth in the healthcare sector, food production and agriculture. This work explores the effect of subinhibitory concentrations of four commonly used biocides (ethanol, hydrogen peroxide, chlorhexidine digluconate...... was determined by an analysis of the transcriptional response of the promoter upstream of tet(M) using β-glucuronidase reporter assays.Results Ethanol significantly (P = 0.01) increased the transfer of Tn916 by 5-fold, whereas hydrogen peroxide, chlorhexidine digluconate and sodium hypochlorite did...

  16. Nano antibody therapy for cancer

    International Nuclear Information System (INIS)

    Venkatachallam, M.; Sivakumar, T.; Nazeema; Venkateswari, P.

    2011-01-01

    Nanomedicine, an offshoot of nanotechnology, refers to highly specific medical intervention at the molecular scale for curing disease or repairing damaged tissues, such as bone, muscle, or nerve. Nanotechnology can have an early, paradigm-changing impact on how clinicians will detect cancer in its earliest stages. Exquisitely sensitive devices constructed of nanoscale components-such as nanocantilevers, nanowires and nanochannels-offer the potential for detecting even the rarest molecular signals associated with malignancy. One of the most pressing needs in clinical oncology is for imaging agents that can identify tumors that are far smaller than is possible with today's technology, at a scale of 100,000 cells rather than 1,000,000,000 cells. A new approach in nanotechnology for treating cancer incorporates nano iron particles and attaches them to an antibody that has targets only cancer cells and not healthy cells. The treatment works in two steps. This treatment is an ingenious way to make localized tumor ablation a systemic treatment. The advantages are incredible. There are absolutely no side effects from this treatment. It is not painful or even uncomfortable. The iron particles get flushed harmlessly from the body. It is not a drug and so the cancer cannot build up a resistance to the treatment. It is a systematic treatment; even cancer cells and tumors that are not known about get heated up and ablated. This treatment can even be used to enhance imaging of the cancer because once the cancer cells are coated with the iron particles, they are easy to identify. Everything depends on how reliably the antibodies target cancer cells and not healthy cells. When used in conjunction with other systemic treatments, such as vaccine treatments, we could be looking at a time when even advanced cancers can be brought under control. (author)

  17. Antibody Production with Synthetic Peptides.

    Science.gov (United States)

    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.

  18. Maturation Pathways of Cross-Reactive HIV-1 Neutralizing Antibodies

    Directory of Open Access Journals (Sweden)

    Dimiter S. Dimitrov

    2009-11-01

    Full Text Available Several human monoclonal antibodies (hmAbs and antibody fragments, including the best characterized in terms of structure-function b12 and Fab X5, exhibit relatively potent and broad HIV-1 neutralizing activity. However, the elicitation of b12 or b12-like antibodies in vivo by vaccine immunogens based on the HIV-1 envelope glycoprotein (Env has not been successful. B12 is highly divergent from the closest corresponding germline antibody while X5 is less divergent. We have hypothesized that the relatively high degree of specific somatic hypermutations may preclude binding of the HIV-1 envelope glycoprotein (Env to closest germline antibodies, and that identifying antibodies that are intermediates in the pathways to maturation could help design novel vaccine immunogens to guide the immune system for their enhanced elicitation. In support of this hypothesis we have previously found that a germline-like b12 (monovalent and bivalent scFv as an Fc fusion protein or IgG lacks measurable binding to an Env as measured by ELISA with a sensitivity in the μM range [1]; here we present evidence confirming and expanding these findings for a panel of Envs. In contrast, a germline-like scFv X5 bound Env with high (nM affinity. To begin to explore the maturation pathways of these antibodies we identified several possible b12 intermediate antibodies and tested their neutralizing activity. These intermediate antibodies neutralized only some HIV-1 isolates and with relatively weak potency. In contrast, germline-like scFv X5 neutralized a subset of the tested HIV-1 isolates with comparable efficiencies to that of the mature X5. These results could help explain the relatively high immunogenicity of the coreceptor binding site on gp120 and the abundance of CD4-induced (CD4i antibodies in HIV-1-infected patients (X5 is a CD4i antibody as well as the maturation pathway of X5. They also can help identify antigens that can bind specifically to b12 germline and

  19. Comparison of cardiac TnI outliers using a contemporary and a high-sensitivity assay on the Abbott Architect platform.

    Science.gov (United States)

    Ryan, J B; Southby, S J; Stuart, L A; Mackay, R; Florkowski, C M; George, P M

    2014-07-01

    Assays for cardiac troponin (cTn) have undergone improvements in sensitivity and precision in recent years. Increased rates of outliers, however, have been reported on various cTn platforms, typically giving irreproducible, falsely higher results. We aimed to evaluate the outlier rate occurring in patients with elevated cTnI using a contemporary and high-sensitivity assay. All patients with elevated cTnI (up to 300 ng/L) performed over a 21-month period were assayed in duplicate. A contemporary assay (Abbott STAT Troponin-I) was used for the first part of the study and subsequently a high-sensitivity assay (Abbott STAT High-Sensitive Troponin-I) was used. Outliers exceeded a calculated critical difference (CD) (CD = z × √2 × SDAnalytical) where z = 3.5 (for probability of 0.0005) and critical outliers also were on a different side of the decision level. The respective outlier and critical outlier rates were 0.22% and 0.10% for the contemporary assay (n = 4009) and 0.18% and 0.13% for the high-sensitivity assay (n = 3878). There was no significant reduction in outlier rate between the two assays (χ(2) = 0.034, P = 0.854). Fifty-six percent of outliers occurred in samples where cTn was an 'add-on' test (and was stored and refrigerated prior to assay). Despite recent improvements in cTn methods, outliers (including critical outliers) still occur at a low rate in both a contemporary and high-sensitivity cTnI assay. Laboratory and clinical staff should be aware of this potential analytical error, particularly in samples with suboptimal sample handling such as add-on tests. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

  20. Dynamic Equilibrium of Cardiac Troponin C's Hydrophobic Cleft and Its Modulation by Ca2+Sensitizers and a Ca2+Sensitivity Blunting Phosphomimic, cTnT(T204E).

    Science.gov (United States)

    Schlecht, William; Dong, Wen-Ji

    2017-10-18

    Several studies have suggested that conformational dynamics are important in the regulation of thin filament activation in cardiac troponin C (cTnC); however, little direct evidence has been offered to support these claims. In this study, a dye homodimerization approach is developed and implemented that allows the determination of the dynamic equilibrium between open and closed conformations in cTnC's hydrophobic cleft. Modulation of this equilibrium by Ca 2+ , cardiac troponin I (cTnI), cardiac troponin T (cTnT), Ca 2+ -sensitizers, and a Ca 2+ -desensitizing phosphomimic of cTnT (cTnT(T204E) is characterized. Isolated cTnC contained a small open conformation population in the absence of Ca 2+ that increased significantly upon the addition of saturating levels of Ca 2+ . This suggests that the Ca 2+ -induced activation of thin filament arises from an increase in the probability of hydrophobic cleft opening. The inclusion of cTnI increased the population of open cTnC, and the inclusion of cTnT had the opposite effect. Samples containing Ca 2+ -desensitizing cTnT(T204E) showed a slight but insignificant decrease in open conformation probability compared to samples with cardiac troponin T, wild type [cTnT(wt)], while Ca 2+ sensitizer treated samples generally increased open conformation probability. These findings show that an equilibrium between the open and closed conformations of cTnC's hydrophobic cleft play a significant role in tuning the Ca 2+ sensitivity of the heart.

  1. [Study of anti-idiotype antibodies to human monoclonal antibody].

    Science.gov (United States)

    Harada, R; Takahashi, N; Owaki, I; Kannagi, R; Endo, N; Morita, N; Inoue, M

    1992-02-01

    A human monoclonal antibody, ll-50 (IgM, lambda), was generated, which reacted specifically with a major of glycolipid present in LS174T colon cancer cells. The glycolipid antigen which reacted with the ll-50 antibody was expected to four sugar residues from its TLC mobility, and it was ascertained that the glycolipid antigen which reacted with ll-50 antibody might be Lc4 antigen [Gal beta 1----3 GLcNAc beta 1----3 Gal beta 1----4 Glc beta 1----1 Cer] judging from TLC immunostaining and ELISA when the reactivity of ll-50 antibody was tested using various pure glycolipids in 3-5 sugar residues as an antigen. Sera in patients with malignant disorders and healthy individuals were analyzed by Sandwich assay of immobilized and biotinylated ll-50 antibody. The serum of the Lc4 antigen recognized by ll-50 antibody was significantly higher in patients with malignant disorders than that in healthy individuals (p less than 0.05). Three mouse monoclonal anti-idiotype antibodies, G3, B3 and C5 (all IgG1), were generated by the immunization of BALB/c mice with ll-50 antibody. These anti-idiotype antibodies specifically bound to to human monoclonal antibody, ll-50 and had a significant inhibitory activity towards the binding of ll-50 antibody to the Lc4 antigen. This indicated that these anti-idiotype antibodies, G3, B3, and C5, were paratope-related anti-idiotype antibodies. G3, B3, and C5 were expected to define the nearest idiotope because they could mutually inhibit ll-50 antibody. Sera in patients with malignant disorders and healthy individuals were analyzed by Sandwich assay of immobilized and biotinylated anti-idiotype antibodies, G3, B3, and C5. As to the ll-50 like antibodies defined by C5 (Id-C5+), the mean serum level in patients with malignant disorders was significantly higher than that in healthy individuals (p less than 0.05). As to the ll-50 like antibodies defined by B3 (Id-B3+), the mean serum level in patients with malignant disorders was significantly higher

  2. A novel detergent-stable solvent-tolerant serine thiol alkaline protease from Streptomyces koyangensis TN650.

    Science.gov (United States)

    Ben Elhoul, Mouna; Zaraî Jaouadi, Nadia; Rekik, Hatem; Bejar, Wacim; Boulkour Touioui, Souraya; Hmidi, Maher; Badis, Abdelmalek; Bejar, Samir; Jaouadi, Bassem

    2015-08-01

    An alkaline proteinase (STAP) was produced from strain TN650 isolated from a Tunisian off-shore oil field and assigned as Streptomyces koyangensis strain TN650 based on physiological and biochemical properties and 16S rRNA gene sequencing. Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis revealed that the purified enzyme was a monomer with a molecular mass of 45125.17-Da. The enzyme had an NH2-terminal sequence of TQSNPPSWGLDRIDQTTAFTKACSIKY, thus sharing high homology with those of Streptomyces proteases. The results showed that this protease was completely inhibited by phenylmethanesulfonyl fluoride (PMSF), diiodopropyl fluorophosphates (DFP), and partially inhibited by 5,5-dithio-bis-(2-nitro benzoic acid) (DTNB), which strongly suggested its belonging to the serine thiol protease family. Using casein as a substrate, the optimum pH and temperature values for protease activity were pH 10 and 70 °C, respectively. The protease was stable at pH 7-10 and 30-60 °C for 24 h. STAP exhibited high catalytic efficiency, significant detergent stability, and elevated organic solvent resistance compared to the SG-XIV proteases from S. griseus and KERAB from Streptomyces sp. AB1. The stap gene encoding STAP was isolated, and its DNA sequence was determined. These properties make STAP a potential candidate for future application in detergent formulations and non-aqueous peptide biocatalysis. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Structural Comparison of Different Antibodies Interacting with Parvovirus Capsids

    Energy Technology Data Exchange (ETDEWEB)

    Hafenstein, Susan; Bowman, Valorie D.; Sun, Tao; Nelson, Christian D.S.; Palermo, Laura M.; Chipman, Paul R.; Battisti, Anthony J.; Parrish, Colin R.; Rossmann, Michael G.; Cornell; Purdue

    2009-05-13

    The structures of canine parvovirus (CPV) and feline parvovirus (FPV) complexed with antibody fragments from eight different neutralizing monoclonal antibodies were determined by cryo-electron microscopy (cryoEM) reconstruction to resolutions varying from 8.5 to 18 {angstrom}. The crystal structure of one of the Fab molecules and the sequence of the variable domain for each of the Fab molecules have been determined. The structures of Fab fragments not determined crystallographically were predicted by homology modeling according to the amino acid sequence. Fitting of the Fab and virus structures into the cryoEM densities identified the footprints of each antibody on the viral surface. As anticipated from earlier analyses, the Fab binding sites are directed to two epitopes, A and B. The A site is on an exposed part of the surface near an icosahedral threefold axis, whereas the B site is about equidistant from the surrounding five-, three-, and twofold axes. One antibody directed to the A site binds CPV but not FPV. Two of the antibodies directed to the B site neutralize the virus as Fab fragments. The differences in antibody properties have been linked to the amino acids within the antibody footprints, the position of the binding site relative to the icosahedral symmetry elements, and the orientation of the Fab structure relative to the surface of the virus. Most of the exposed surface area was antigenic, although each of the antibodies had a common area of overlap that coincided with the positions of the previously mapped escape mutations.

  4. Radioimmunodetection of human melanoma tumor xenografts with human monoclonal antibodies

    International Nuclear Information System (INIS)

    Gomibuchi, Makoto; Saxton, R.E.; Lake, R.R.; Katano, Mitsuo; Irie, R.F.

    1986-01-01

    A human IgM monoclonal antibody has been established that defines a tumor-associated membrane antigen expressed on human melanoma cells. The antigen has been identified as the ganglioside GD2. In this paper, the authors describe the potential usefulness of the human monoclonal antibody for radioimaging. Nude mice bearing tumors derived from a human melanoma cell line were used as a model. Antibody activity was degradated significantly after labeling with 131 I by the use of a modified chloramine-T method. After testing various concentrations, labeled antibody of a specific activity of 2.8μCi/μg produced the best results. Balb/c nude mice bearing a GD2-positive M14 melanoma cell line were injected with 10-30μg of labeled antibody, and its radiolocalization in different organs and in the whole body were evaluated. The best tumor image was obtained on Day 6. The labeled antibody uptake ratio between tumor and muscle was 9.2:1; the ratio between tumor and liver was 1.4:1. These studies represent the first report of experimental tumor imaging with human monoclonal antibody. Human monoclonals will probably prove to be superior reagents for tumor imaging in melanoma patients if the problem of anti-body radiolysis is resolved. (author)

  5. Seroprevalence Survey of Rubella Antibodies among Pregnant ...

    African Journals Online (AJOL)

    Key words: Rubella virus, teratogen, antibodies, Maiduguri. La rubéole est une infection virale évitable par la vaccination. Son agent étiologique, virus de la rubéole a été identifié comme un tératogène humain capable de provoquer le spectre de malformation congénitale décrite comme le syndrome de rubéole congénitale ...

  6. Antibody induction therapy for lung transplant recipients

    DEFF Research Database (Denmark)

    Penninga, Luit; Møller, Christian H; Penninga, Ida Elisabeth Irene

    2013-01-01

    Lung transplantation has become a valuable and well-accepted treatment option for most end-stage lung diseases. Lung transplant recipients are at risk of transplanted organ rejection, and life-long immunosuppression is necessary. Clear evidence is essential to identify an optimal, safe...... and effective immunosuppressive treatment strategy for lung transplant recipients. Consensus has not yet been achieved concerning use of immunosuppressive antibodies against T-cells for induction following lung transplantation....

  7. Detection of novel diagnostic antibodies in ankylosing spondylitis: An overview.

    Science.gov (United States)

    Quaden, Dana H F; De Winter, Liesbeth M; Somers, Veerle

    2016-08-01

    Ankylosing spondylitis (AS) is a debilitating, chronic, rheumatic disease characterized by inflammation and new bone formation resulting in fusion of the spine and sacroiliac joints. Since early treatment is impeded by a delayed diagnosis, it is highly important to find new biomarkers that improve early diagnosis and may also contribute to a better assessment of disease activity, prognosis and therapy response in AS. Because of the absence of rheumatoid factor, AS was long assumed to have a seronegative character and antibodies are thus not considered a hallmark of the disease. However, emerging evidence suggests plasma cells and autoantibodies to be involved in the disease course. In this review, the role of B cells and antibodies in AS is discussed. Furthermore, an overview is provided of antibodies identified in AS up till now, and their diagnostic potential. Many of these antibody responses were based on small study populations and further validation is lacking. Moreover, most were identified by a hypothesis-driven approach and thus limited to antibodies against targets that are already known to be involved in AS pathogenesis. Hence, we propose an unbiased approach to identify novel diagnostic antibodies. The already successfully applied techniques cDNA phage display and serological antigen selection will be used to identify antibodies against both known and new antigen targets in AS plasma. These newly identified antibodies will enhance early diagnosis of AS and provide more insight into the underlying disease pathology, resulting in a more effective treatment strategy and eventually an improved disease outcome. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. The antibody Hijikata Tatsumi

    Directory of Open Access Journals (Sweden)

    Éden Peretta

    2012-11-01

    Full Text Available Considered one of the most influential modern dance representatives in Japan, Tatsumi Hijikata’s work was a milestone in the Japanese post-war experimental artistic scene. Heretic son of his time, he staged a fertile mix of artistic and cultural influences, overlapping subversive elements of European arts and philosophy with radical references from pre-modern Japanese culture. In this way he built the foundations of its unstable antibody, its political-artistic project of dissolution of a organism, both physical and social.

  9. The emergence of antibody therapies for Ebola.

    Science.gov (United States)

    Hiatt, Andrew; Pauly, Michael; Whaley, Kevin; Qiu, Xiangguo; Kobinger, Gary; Zeitlin, Larry

    2015-12-23

    This review describes the history of Ebola monoclonal antibody (mAb) development leading up to the recent severe Ebola outbreak in West Africa. The Ebola virus has presented numerous perplexing challenges in the long effort to develop therapeutic antibody strategies. Since the first report of a neutralizing human anti-Ebola mAb in 1999, the straightforward progression from in vitro neutralization resulting in in vivo protection and therapy has not occurred. A number of mAbs, including the first reported, failed to protect non-human primates (NHPs) in spite of protection in rodents. An appreciation of the role of effector functions to antibody efficacy has contributed significantly to understanding mechanisms of in vivo protection. However a crucial contribution, as measured by post-exposure therapy of NHPs, involved the comprehensive testing of mAb cocktails. This effort was aided by the use of plant production technology where various combinations of mAbs could be rapidly produced and tested. Introduction of appropriate modifications, such as specific glycan profiles, also improved therapeutic efficacy. The resulting cocktail, ZMapp™, consists of three mAbs that were identified from numerous mAb candidates. ZMapp™ \\ is now being evaluated in human clinical trials but has already played a role in bringing awareness to the potential of antibody therapy for Ebola.

  10. Developing recombinant antibodies for biomarker detection

    Energy Technology Data Exchange (ETDEWEB)

    Baird, Cheryl L.; Fischer, Christopher J.; Pefaur, Noah B.; Miller, Keith D.; Kagen, Jacob; Srivastava, Sudhir; Rodland, Karin D.

    2010-10-01

    Monoclonal antibodies (mAbs) have an essential role in biomarker validation and diagnostic assays. A barrier to pursuing these applications is the reliance on immunization and hybridomas to produce mAbs, which is time-consuming and may not yield the desired mAb. We recommend a process flow for affinity reagent production that utilizes combinatorial protein display systems (eg, yeast surface display or phage display) rather than hybridomas. These systems link a selectable phenotype-binding conferred by an antibody fragment-with a means for recovering the encoding gene. Recombinant libraries obtained from immunizations can produce high-affinity antibodies (<10 nM) more quickly than other methods. Non-immune libraries provide an alternate route when immunizations are not possible, or when suitable mAbs are not recovered from an immune library. Directed molecular evolution (DME) is an integral part of optimizing mAbs obtained from combinatorial protein display, but can also be used on hybridoma-derived mAbs. Variants can easily be obtained and screened to increase the affinity of the parent mAb (affinity maturation). We discuss examples where DME has been used to tailor affinity reagents to specific applications. Combinatorial protein display also provides an accessible method for identifying antibody pairs, which are necessary for sandwich-type diagnostic assays.

  11. Single Domain Antibodies as New Biomarker Detectors

    Science.gov (United States)

    Fischer, Katja; Leow, Chiuan Yee; Chuah, Candy; McCarthy, James

    2017-01-01

    Biomarkers are defined as indicators of biological processes, pathogenic processes, or pharmacological responses to a therapeutic intervention. Biomarkers have been widely used for early detection, prediction of response after treatment, and for monitoring the progression of diseases. Antibodies represent promising tools for recognition of biomarkers, and are widely deployed as analytical tools in clinical settings. For immunodiagnostics, antibodies are now exploited as binders for antigens of interest across a range of platforms. More recently, the discovery of antibody surface display and combinatorial chemistry techniques has allowed the exploration of new binders from a range of animals, for instance variable domains of new antigen receptors (VNAR) from shark and variable heavy chain domains (VHH) or nanobodies from camelids. These single domain antibodies (sdAbs) have some advantages over conventional murine immunoglobulin owing to the lack of a light chain, making them the smallest natural biomarker binders thus far identified. In this review, we will discuss several biomarkers used as a means to validate diseases progress. The potential functionality of modern singe domain antigen binders derived from phylogenetically early animals as new biomarker detectors for current diagnostic and research platforms development will be described. PMID:29039819

  12. Single Domain Antibodies as New Biomarker Detectors

    Directory of Open Access Journals (Sweden)

    Chiuan Herng Leow

    2017-10-01

    Full Text Available Biomarkers are defined as indicators of biological processes, pathogenic processes, or pharmacological responses to a therapeutic intervention. Biomarkers have been widely used for early detection, prediction of response after treatment, and for monitoring the progression of diseases. Antibodies represent promising tools for recognition of biomarkers, and are widely deployed as analytical tools in clinical settings. For immunodiagnostics, antibodies are now exploited as binders for antigens of interest across a range of platforms. More recently, the discovery of antibody surface display and combinatorial chemistry techniques has allowed the exploration of new binders from a range of animals, for instance variable domains of new antigen receptors (VNAR from shark and variable heavy chain domains (VHH or nanobodies from camelids. These single domain antibodies (sdAbs have some advantages over conventional murine immunoglobulin owing to the lack of a light chain, making them the smallest natural biomarker binders thus far identified. In this review, we will discuss several biomarkers used as a means to validate diseases progress. The potential functionality of modern singe domain antigen binders derived from phylogenetically early animals as new biomarker detectors for current diagnostic and research platforms development will be described.

  13. Antibodies to actin in autoimmune haemolytic anaemia

    Directory of Open Access Journals (Sweden)

    Ritzmann Mathias

    2010-03-01

    Full Text Available Abstract Background In autoimmune haemolytic anaemia (AIHA, autoreactive antibodies directed against red blood cells are up-regulated, leading to erythrocyte death. Mycoplasma suis infections in pigs induce AIHA of both the warm and cold types. The aim of this study was to identify the target autoantigens of warm autoreactive IgG antibodies. Sera from experimentally M. suis-infected pigs were screened for autoreactivity. Results Actin-reactive antibodies were found in the sera of 95% of all animals tested. The reactivity was species-specific, i.e. reactivity with porcine actin was significantly higher than with rabbit actin. Sera of animals previously immunised with the M. suis adhesion protein MSG1 showed reactivity with actin prior to infection with M. suis indicating that molecular mimicry is involved in the specific autoreactive mechanism. A potentially cross-reactive epitope was detected. Conclusions This is the first report of autoreactive anti-actin antibodies involved in the pathogenesis of autoimmune haemolytic anaemia.

  14. Antibody Maturation in Trypanosoma cruzi-Infected Rats

    Science.gov (United States)

    Marcipar, Iván S.; Risso, Marikena G.; Silber, Ariel M.; Revelli, Silvia; Marcipar, Alberto J.

    2001-01-01

    The study of antibody avidity changes during infection has improved the understanding of the pathologic processes involved in several infectious diseases. In some infections, like toxoplasmosis, this information is being used for diagnostic purposes. Results of the evolution of antibody avidity for different specific antigens in Trypanosome cruzi-infected rats are presented. A Western blotting technique, combined with avidity analysis to identify antigens that elicit high-avidity antibodies, is suggested. In this system, antibodies showed high avidity values only during the chronic phase of infection and only in relation to antibodies against 21-, 33-, 41-, 42-, 56-, 58-, 66-, and 72-kDa antigens. Finally, a 97-kDa T. cruzi antigen, which was recognized by high-avidity antibodies and occurred in noninfected rats, was identified. These results allow us to evaluate the different antigens in chagasic infection. Our results show that with the correct choice of antigen it is possible to detect differences in maturation of antibodies and to discriminate, in an experimental model, between recent (acute) and chronic infections. PMID:11427430

  15. Cooperativity Enables Non-neutralizing Antibodies to Neutralize Ebolavirus

    Directory of Open Access Journals (Sweden)

    Katie A. Howell

    2017-04-01

    Full Text Available Drug combinations are synergistic when their combined efficacy exceeds the sum of the individual actions, but they rarely include ineffective drugs that become effective only in combination. We identified several “enabling pairs” of neutralizing and non-neutralizing anti-ebolavirus monoclonal antibodies, whose combination exhibited new functional profiles, including transforming a non-neutralizing antibody to a neutralizer. Sub-neutralizing concentrations of antibodies 2G4 or m8C4 enabled non-neutralizing antibody FVM09 (IC50 >1 μM to exhibit potent neutralization (IC50 1–10 nM. While FVM09 or m8C4 alone failed to protect Ebola-virus-infected mice, a combination of the two antibodies provided 100% protection. Furthermore, non-neutralizers FVM09 and FVM02 exponentially enhanced the potency of two neutralizing antibodies against both Ebola and Sudan viruses. We identified a hotspot for the binding of these enabling antibody pairs near the interface of the glycan cap and GP2. Enabling cooperativity may be an underappreciated phenomenon for viruses, with implications for the design and development of immunotherapeutics and vaccines.

  16. The future of monoclonal antibody technology

    OpenAIRE

    Zider, Alexander; Drakeman, Donald L

    2010-01-01

    With the rapid growth of monoclonal antibody-based products, new technologies have emerged for creating modified forms of antibodies, including fragments, conjugates and multi-specific antibodies. We created a database of 450 therapeutic antibodies in development to determine which technologies and indications will constitute the “next generation” of antibody products. We conclude that the antibodies of the future will closely resemble the antibodies that have already been approved for commer...

  17. Discovery Of Human Antibodies Against Spitting Cobra Toxins

    DEFF Research Database (Denmark)

    Bojsen-Møller, Laura; Lohse, Brian; Harrison, Robert

    Current snakebite envenoming treatment options consist of animal-derived antisera and are associated with severe adverse reactions due to the heterologous nature of the animal-derived antibodies present in these antisera, and the presence of therapeutically irrelevant antibodies. The African...... spitting cobras are among the most medically important snakes in sub-Saharan regions due to the severity of the clinical outcomes caused by their cytotoxic venom, which is derived from cytotoxins of the 3FTx toxin family and PLA2. Here we report the results of our progress in identifying human antibodies...... targeting relevant toxins from the venom of the black necked spitting cobra (Naja nigricolis)....

  18. Quantitative assessment of antibody internalization with novel monoclonal antibodies against Alexa fluorophores.

    Directory of Open Access Journals (Sweden)

    Sindy Liao-Chan

    Full Text Available Antibodies against cell surface antigens may be internalized through their specific interactions with these proteins and in some cases may induce or perturb antigen internalization. The anti-cancer efficacy of antibody-drug conjugates is thought to rely on their uptake by cancer cells expressing the surface antigen. Numerous techniques, including microscopy and flow cytometry, have been used to identify antibodies with desired cellular uptake rates. To enable quantitative measurements of internalization of labeled antibodies, an assay based on internalized and quenched fluorescence was developed. For this approach, we generated novel anti-Alexa Fluor monoclonal antibodies (mAbs that effectively and specifically quench cell surface-bound Alexa Fluor 488 or Alexa Fluor 594 fluorescence. Utilizing Alexa Fluor-labeled mAbs against the EphA2 receptor tyrosine kinase, we showed that the anti-Alexa Fluor reagents could be used to monitor internalization quantitatively over time. The anti-Alexa Fluor mAbs were also validated in a proof of concept dual-label internalization assay with simultaneous exposure of cells to two different mAbs. Importantly, the unique anti-Alexa Fluor mAbs described here may also enable other single- and dual-label experiments, including label detection and signal enhancement in macromolecules, trafficking of proteins and microorganisms, and cell migration and morphology.

  19. Tabhu: tools for antibody humanization

    DEFF Research Database (Denmark)

    Olimpieri, Pier Paolo; Marcatili, Paolo; Tramontano, Anna

    2015-01-01

    and time-consuming experiments. Here we present tools for antibody humanization (Tabhu) a web server for antibody humanization. Tabhu includes tools for human template selection, grafting, back-mutation evaluation, antibody modelling and structural analysis, helping the user in all the critical steps...... elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity...... of the humanization experiment protocol....

  20. Anticardiolipin antibody and anti-beta 2 glycoprotein I antibody assays.

    Science.gov (United States)

    Raby, Anne; Moffat, Karen; Crowther, Mark

    2013-01-01

    Antiphospholipid syndrome (APS) is an autoimmune disease and is a risk factor for a number of clinical manifestations; the classic presentations include fetal death or thrombosis (arterial or venous thromboembolism), in the presence of persistently increased titers of antiphospholipid (aPL) antibodies. The actual cause of APS is unknown but thought to be multifactorial. The disease is characterized by the presence of a heterogenous population of autoantibodies against phospholipid-binding proteins. APS presents either in isolation with no evidence of an underlying disease or in concert with an autoimmune disease such as systemic lupus erythematosus or rheumatoid arthritis. The wide diversity in clinical presentation often causes difficulty in identifying and treating patients and therefore a concise laboratory report containing interpretative comments is required to provide needed guidance to the clinician. For a diagnosis of APS to be made both clinical and laboratory classification criteria must be met. Laboratory testing to identify aPL antibodies includes lupus anticoagulant (liquid-based clotting assays) and immunological solid-phase assays (usually enzyme-linked immunosorbent assay formats) for IgG and/or IgM anticardiolipin (aCL) antibodies and anti-beta 2 glycoprotein I (β2-GPI) antibodies. Other autoantibodies, such as those directed against anionic phospholipids, can also be assayed; however they are not of clinical significance. Participation in a quality assurance program and an in-depth technical and clinical understanding of testing for aPL antibodies are required, as methods are limited by poor robustness, reproducibility, specificity, and standardization. Testing is further complicated by the lack of a "gold standard" laboratory test to diagnose or classify a patient as having APS. This chapter discusses the clinical and laboratory theoretical and technical aspects of aCL and anti-β2GPI antibody assays.

  1. Lupus Anticoagulant and Anticardiolipin Antibodies in Unexplained Fetal Losses

    OpenAIRE

    ALPER, Gülinnaz

    2014-01-01

    Lupus anticoagulant (LA) and anti-cardiolipin antibodies (ACAs) are acquired antiphospholipid antibodies (APAs), which are considered to be important markers for pregnancy losses and intrauterine fetal demise. LA and ACAs have anticoagulant effects in vitro and thrombotic effects in vivo and are considered to be the cause of recur-rent pregnancy losses (RPLs), resulting from placental vascular thrombosis and infarction. The aim of this study was to identify the most sensitive and specific met...

  2. Antigen-antibody reactions of UV-irradiated phage DNA

    International Nuclear Information System (INIS)

    Fink, A.

    1976-01-01

    The observation of others could be confirmed that UV-irradiated DNA is a better immunogen than unirradiated DNA. The author's immune sera contained a high amount of antibodies with a specific action against photoproducts in the DNA. The thymine dimer was identified as relevant photoproduct and thus as antigenic determinant. In comparison, the amount of unspecific antibodies reacting with denaturated DNA was low and varied between sera. Thymin-dimer antibodies showed a high specificity without cross-reaction with other pyrimidine dimers such as anti CC and anti CT; they belong to the class of IgG molecules. UV-irradiated dinucleotide dTpT is sufficient to induce the formation of antibodies reacting with the cis-syn thymine dimers in UV-irradiated DNA. Antibody binding is proportional to the UV doses applied to the DNA. When using completely denaturated DNA, there is a linear increase changing into a plateau at higher doses. The extent of antigen-antibody binding is strongly dependent on the degree of denaturation of the DNA. With increasing denaturation, the antibody binding of the DNA increases. The antigen-antibody reaction can thus be used to estimate the degree of denaturation of the DNA. There were no signs of an influence of the degree of denaturation of the DNA on the quantum yield of thymine dimers. The different amounts of antibodies is therefore due to the masking of thymine dimers in native DNA. When irradiating intact phage particles, there was no sign of an influence of the phages' protein covers on the antibody binding capacity of DNA compared with DNA irradiated in vitro. (orig.) [de

  3. Theranostics Using Antibodies and Antibody-Related Therapeutics

    NARCIS (Netherlands)

    Moek, Kirsten L; Giesen, Danique; Kok, Iris C; de Groot, Derk Jan A; Jalving, Mathilde; Fehrmann, Rudolf S N; Lub-de Hooge, Marjolijn N; Brouwers, Adrienne H; de Vries, Elisabeth G E

    In theranostics, radiolabeled compounds are used to determine a treatment strategy by combining therapeutics and diagnostics in the same agent. Monoclonal antibodies (mAbs) and antibody-related therapeutics represent a rapidly expanding group of cancer medicines. Theranostic approaches using these

  4. Discovery and characterization of antibody variants using mass spectrometry-based comparative analysis for biosimilar candidates of monoclonal antibody drugs.

    Science.gov (United States)

    Li, Wenhua; Yang, Bin; Zhou, Dongmei; Xu, Jun; Ke, Zhi; Suen, Wen-Chen

    2016-07-01

    Liquid chromatography mass spectrometry (LC-MS) is the most commonly used technique for the characterization of antibody variants. MAb-X and mAb-Y are two approved IgG1 subtype monoclonal antibody drugs recombinantly produced in Chinese hamster ovary (CHO) cells. We report here that two unexpected and rare antibody variants have been discovered during cell culture process development of biosimilars for these two approved drugs through intact mass analysis. We then used comprehensive mass spectrometry-based comparative analysis including reduced light, heavy chains, and domain-specific mass as well as peptide mapping analysis to fully characterize the observed antibody variants. The "middle-up" mass comparative analysis demonstrated that the antibody variant from mAb-X biosimilar candidate was caused by mass variation of antibody crystalline fragment (Fc), whereas a different variant with mass variation in antibody antigen-binding fragment (Fab) from mAb-Y biosimilar candidate was identified. Endoproteinase Lys-C digested peptide mapping and tandem mass spectrometry analysis further revealed that a leucine to glutamine change in N-terminal 402 site of heavy chain was responsible for the generation of mAb-X antibody variant. Lys-C and trypsin coupled non-reduced and reduced peptide mapping comparative analysis showed that the formation of the light-heavy interchain trisulfide bond resulted in the mAb-Y antibody variant. These two cases confirmed that mass spectrometry-based comparative analysis plays a critical role for the characterization of monoclonal antibody variants, and biosimilar developers should start with a comprehensive structural assessment and comparative analysis to decrease the risk of the process development for biosimilars. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Some transition metal ions complexes of tricine (Tn and amino acids: pH-titration, synthesis and antimicrobial activity

    Directory of Open Access Journals (Sweden)

    M.E. Zayed

    2014-12-01

    Full Text Available Equilibrium studies have been carried out on complex formation of M(II (M = Co(II, Cu(II and Zn(II with tricine (Tn and L = amino acids in aqueous solution, at 25 °C and ionic strength of I = 0.1 M (NaNO3. The ternary complexes of amino acids are formed by simultaneous reactions. The concentration distribution of the complexes is evaluated. The solid complexes of [M(II–Tn–Histidine (Hist] have been synthesized and characterized by elemental analysis, infrared, magnetic and conductance measurements. The synthesized complexes have been screened for their antibacterial activities and the complexes show a significant antibacterial activity against four bacterial species: Staphylococcus aureus (Gram +ve, Streptococcus pyogenesr (Gram +ve, Serratia marcescens (Gram −ve and Escherichia coli (Gram −ve. The activity increases by increasing the concentration of the complexes.

  6. Tn5090-like class 1 integron carrying bla(VIM-2) in a Pseudomonas putida strain from Portugal.

    Science.gov (United States)

    Santos, C; Caetano, T; Ferreira, S; Mendo, S

    2010-10-01

    Three Pseudomonas putida strains containing bla(VIM-2) were isolated from an inanimate surface of a female ward sanitary facility in the Hospital Infante D. Pedro, Aveiro. A novel class 1 integron was found in strain Pp2 (aacA4/bla(VIM-2)/aac6'-IIc disrupted by an insertion sequence IS1382), and strain Pp1 was found to carry a class 1 integron (aacA7/bla(VIM-2)/aacC1/aacA4), which is described for the first time in this species. Strain PF1 carries a class 1 integron associated with a Tn5090-like transposon, constituting the first finding of this type of arrangement in a strain from Portugal. This association highlights further dissemination of bla(VIM-2) in environmental hospital isolates. © 2010 The Authors. Journal Compilation © 2010 European Society of Clinical Microbiology and Infectious Diseases.

  7. Antibodies and Plasmodium falciparum merozoites

    NARCIS (Netherlands)

    Ramasamy, R; Ramasamy, M; Yasawardena, S

    There is considerable interest in using merozoite proteins in a vaccine against falciparum malaria. Observations that antibodies to merozoite surface proteins block invasion are a basis for optimism. This article draws attention to important and varied aspects of how antibodies to Plasmodium

  8. Catalytic Antibodies: Concept and Promise

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 12; Issue 11. Catalytic Antibodies: Concept and Promise. Desirazu N Rao Bharath Wootla. General Article Volume 12 Issue ... Keywords. Catalytic antibodies; abzymes; hybridome technology; Diels– Alder reaction; Michaelis– Menten kinetics; Factor VIII.

  9. Antiphospholipid antibodies: standardization and testing.

    Science.gov (United States)

    Riley, R S; Friedline, J; Rogers, J S

    1997-09-01

    A phenomenon originally scorned as a laboratory nuisance has turned out to be an important cause of thromboembolism, fetal death, and other forms of human disease. Investigations of this inaptly named "lupus anticoagulant" has led to the discovery of at least two distinct types of autoimmune antibodies. In spite of recent discoveries regarding the pathophysiology of these antibodies, their clinical significance is still controversial.

  10. Educational paper: Primary antibody deficiencies

    NARCIS (Netherlands)

    G.J.A. Driessen (Gertjan); M. van der Burg (Mirjam)

    2011-01-01

    textabstractPrimary antibody deficiencies (PADs) are the most common primary immunodeficiencies and are characterized by a defect in the production of normal amounts of antigen-specific antibodies. PADs represent a heterogeneous spectrum of conditions, ranging from often asymptomatic selective IgA

  11. Annual Performance Evaluation of a Pair of Energy Efficient Houses (WC3 and WC4) in Oak Ridge, TN

    Energy Technology Data Exchange (ETDEWEB)

    Biswas, Kaushik [ORNL; Christian, Jeffrey E [ORNL; Gehl, Anthony C [ORNL; Jackson, Roderick K [ORNL; Boudreaux, Philip R [ORNL

    2012-04-01

    Beginning in 2008, two pairs of energy-saver houses were built at Wolf Creek in Oak Ridge, TN. These houses were designed to maximize energy efficiency using new ultra-high-efficiency components emerging from ORNL s Cooperative Research and Development Agreement (CRADA) partners and others. The first two houses contained 3713 square feet of conditioned area and were designated as WC1 and WC2; the second pair consisted of 2721 square feet conditioned area with crawlspace foundation and they re called WC3 and WC4. This report is focused on the annual energy performance of WC3 and WC4, and how they compare against a previously benchmarked maximum energy efficient house of a similar footprint. WC3 and WC4 are both about 55-60% more efficient than traditional new construction. Each house showcases a different envelope system: WC3 is built with advanced framing featured cellulose insulation partially mixed with phase change materials (PCM); and WC4 house has cladding composed of an exterior insulation and finish system (EIFS). The previously benchmarked house was one of three built at the Campbell Creek subdivision in Knoxville, TN. This house (CC3) was designed as a transformation of a builder house (CC1) with the most advanced energy-efficiency features, including solar electricity and hot water, which market conditions are likely to permit within the 2012 2015 period. The builder house itself was representative of a standard, IECC 2006 code-certified, all-electric house built by the builder to sell around 2005 2008.

  12. [Antibody induction after intrauterine interventions].

    Science.gov (United States)

    Hoch, J; Giers, G; Bald, R; Hansmann, M; Hanfland, P

    1993-06-01

    Immunohematologic and clinical data, i.e., antibody profile, location of the placenta, mode of cordocentesis, obtained from 48 pregnant patients with irregular erythrocyte antibodies during the last 2 years have been retrospectively evaluated. All fetuses of the patients received intrauterine transfusions for the treatment of fetal erythroblastosis. In 16 (33%) patients (group I) a secondarily induced antibody was detected after the onset of intrauterine transfusion therapy. 32 (67%) patients (group II) did not further develop new antibody specificities. Group I exhibited a significantly different distribution in the location of the placenta (p pregnant women. In group I a 5-fold higher rate of anterior than posterior placenta location was found. The mode of cordocentesis differed significantly (p antibodies by invasive intrauterine interventions in our patients depended indirectly on the location of the placenta and directly on the mode of the puncture (trans- vs. paraplacental access).

  13. Recent developments in monoclonal antibody radiolabeling techniques

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, S.C.; Mease, R.C.

    1989-01-01

    Monoclonal antibodies (MAbs) have shown the potential to serve as selective carriers of radionuclides to specific in vivo antigens. Accordingly, there has been an intense surge of research activity in an effort to develop and evaluate MAb-based radiopharmaceuticals for tumor imaging (radioimmunoscintigraphy) and therapy (radioimmunotherapy), as well as for diagnosing nonmalignant diseases. A number of problems have recently been identified, related to the MAbs themselves and to radiolabeling techniques, that comprise both the selectivity and the specificity of the in vivo distribution of radiolabeled MAbs. This paper will address some of these issues and primarily discuss recent developments in the techniques for radiolabeling monoclonal antibodies that may help resolve problems related to the poor in vivo stability of the radiolabel and may thus produce improved biodistribution. Even though many issues are identical with therapeutic radionuclides, the discussion will focus mainly on radioimmunoscintigraphic labels. 78 refs., 6 tabs.

  14. Recent developments in monoclonal antibody radiolabeling techniques

    International Nuclear Information System (INIS)

    Srivastava, S.C.; Mease, R.C.

    1989-01-01

    Monoclonal antibodies (MAbs) have shown the potential to serve as selective carriers of radionuclides to specific in vivo antigens. Accordingly, there has been an intense surge of research activity in an effort to develop and evaluate MAb-based radiopharmaceuticals for tumor imaging (radioimmunoscintigraphy) and therapy (radioimmunotherapy), as well as for diagnosing nonmalignant diseases. A number of problems have recently been identified, related to the MAbs themselves and to radiolabeling techniques, that comprise both the selectivity and the specificity of the in vivo distribution of radiolabeled MAbs. This paper will address some of these issues and primarily discuss recent developments in the techniques for radiolabeling monoclonal antibodies that may help resolve problems related to the poor in vivo stability of the radiolabel and may thus produce improved biodistribution. Even though many issues are identical with therapeutic radionuclides, the discussion will focus mainly on radioimmunoscintigraphic labels. 78 refs., 6 tabs

  15. Integration of Antibody Array Technology into Drug Discovery and Development.

    Science.gov (United States)

    Huang, Wei; Whittaker, Kelly; Zhang, Huihua; Wu, Jian; Zhu, Si-Wei; Huang, Ruo-Pan

    Antibody arrays represent a high-throughput technique that enables the parallel detection of multiple proteins with minimal sample volume requirements. In recent years, antibody arrays have been widely used to identify new biomarkers for disease diagnosis or prognosis. Moreover, many academic research laboratories and commercial biotechnology companies are starting to apply antibody arrays in the field of drug discovery. In this review, some technical aspects of antibody array development and the various platforms currently available will be addressed; however, the main focus will be on the discussion of antibody array technologies and their applications in drug discovery. Aspects of the drug discovery process, including target identification, mechanisms of drug resistance, molecular mechanisms of drug action, drug side effects, and the application in clinical trials and in managing patient care, which have been investigated using antibody arrays in recent literature will be examined and the relevance of this technology in progressing this process will be discussed. Protein profiling with antibody array technology, in addition to other applications, has emerged as a successful, novel approach for drug discovery because of the well-known importance of proteins in cell events and disease development.

  16. Recent Advances in Monoclonal Antibody Therapies for Multiple Sclerosis

    Science.gov (United States)

    Stavropoulos, Nikolaos; Wittenberg, Nathan J.; Dasari, Harika; Abdelrahim, Murtada A.; Henley, John R.; Oh, Sang-Hyun; Warrington, Arthur E.; Rodriguez, Moses

    2016-01-01

    Introduction Multiple sclerosis (MS) is the most common chronic inflammatory, demyelinating disease of the CNS and results in neurological disability. Existing immunomodulatory and immunosuppressive approaches lower the number of relapses but do not cure or reverse existing deficits nor improve long-term disability in MS patients. Areas Covered Monogenic antibodies were described as treatment options for MS, however the immunogenicity of mouse antibodies hampered the efficacy of potential therapeutics in humans. Availability of improved antibody production technologies resulted in a paradigm shift in MS treatment strategies. In this review, an overview of immunotherapies for MS that use conventional monoclonal antibodies reactive to immune system and their properties and mechanisms of action will be discussed, including recent advances in MS therapeutics and highlight natural autoantibodies (NAbs) that directly target CNS cells. Expert Opinion Recent challenges for MS therapy are the identification of relevant molecular and cellular targets, time frame of treatment, and antibody toxicity profiles to identify safe treatment options for MS patients. The application of monoclonal antibody therapies with better biological efficacy associated with minimum side effects possesses huge clinical potential. Advances in monoclonal antibody technologies that directly target cells of nervous system may promote the CNS regeneration field from bench to bedside. PMID:26914737

  17. 76 FR 4147 - Putnam-Cumberland, TN-Improve Power Supply

    Science.gov (United States)

    2011-01-24

    ... environmental impacts of the construction, operation, and maintenance of proposed new and upgraded power... assessment (EA) or an environmental impact statement (EIS) to address the potential environmental effects of... potential environmental impacts and other important issues identified in the scoping process, as well as...

  18. Combining Shigella Tn-seq data with gold-standard E. coli gene deletion data suggests rare transitions between essential and non-essential gene functionality.

    Science.gov (United States)

    Freed, Nikki E; Bumann, Dirk; Silander, Olin K

    2016-09-06

    Gene essentiality - whether or not a gene is necessary for cell growth - is a fundamental component of gene function. It is not well established how quickly gene essentiality can change, as few studies have compared empirical measures of essentiality between closely related organisms. Here we present the results of a Tn-seq experiment designed to detect essential protein coding genes in the bacterial pathogen Shigella flexneri 2a 2457T on a genome-wide scale. Superficial analysis of this data suggested that 481 protein-coding genes in this Shigella strain are critical for robust cellular growth on rich media. Comparison of this set of genes with a gold-standard data set of essential genes in the closely related Escherichia coli K12 BW25113 revealed that an excessive number of genes appeared essential in Shigella but non-essential in E. coli. Importantly, and in converse to this comparison, we found no genes that were essential in E. coli and non-essential in Shigella, implying that many genes were artefactually inferred as essential in Shigella. Controlling for such artefacts resulted in a much smaller set of discrepant genes. Among these, we identified three sets of functionally related genes, two of which have previously been implicated as critical for Shigella growth, but which are dispensable for E. coli growth. The data presented here highlight the small number of protein coding genes for which we have strong evidence that their essentiality status differs between the closely related bacterial taxa E. coli and Shigella. A set of genes involved in acetate utilization provides a canonical example. These results leave open the possibility of developing strain-specific antibiotic treatments targeting such differentially essential genes, but suggest that such opportunities may be rare in closely related bacteria.

  19. Mathematical analysis of dengue virus antibody dynamics

    Science.gov (United States)

    Perera, Sulanie; Perera, SSN

    2018-03-01

    Dengue is a mosquito borne viral disease causing over 390 million infections worldwide per annum. Even though information on how infection is controlled and eradicated from the body is lacking, antibodies are thought to play a major role in clearing the virus. In this paper, a non-linear conceptual dynamical model with humoral immune response and absorption effect has been proposed for primary dengue infection. We have included the absorption of pathogens into uninfected cells since this effect causes the virus density in the blood to decrease. The time delay that arises in the production of antibodies was accounted and is introduced through a continuous function. The basic reproduction number R0 is computed and a detailed stability analysis is done. Three equilibrium states, namely the infection free equilibrium, no immune equilibrium and the endemic equilibrium were identified and the existence and the stability conditions of these steady states were obtained. Numerical simulations proved the results that were obtained. By establishing the characteristic equation of the model at infection free equilibrium, it was observed that the infection free equilibrium is locally asymptotically stable if R0 1. Stability regions are identified for infection free equilibrium state with respect to the external variables and it is observed as the virus burst rate increases, the stability regions would decrease. These results implied that for higher virus burst rates, other conditions in the body must be strong enough to eliminate the disease completely from the host. The effect of time delay of antibody production on virus dynamics is discussed. It was seen that as the time delay in production of antibodies increases, the time for viral decline also increased. Also it was observed that the virus count goes to negligible levels within 7 - 14 days after the onset of symptoms as seen in dengue infections.

  20. Altered Antibody Profiles against Common Infectious Agents in Chronic Disease

    Science.gov (United States)

    Burbelo, Peter D.; Ching, Kathryn H.; Morse, Caryn G.; Alevizos, Ilias; Bayat, Ahmad; Cohen, Jeffrey I.; Ali, Mir A.; Kapoor, Amit; Browne, Sarah K.; Holland, Steven M.; Kovacs, Joseph A.; Iadarola, Michael J.

    2013-01-01

    Despite the important diagnostic value of evaluating antibody responses to individual human pathogens, antibody profiles against multiple infectious agents have not been used to explore health and disease mainly for technical reasons.  We hypothesized that the interplay between infection and chronic disease might be revealed by profiling antibodies against multiple agents. Here, the levels of antibodies against a panel of 13 common infectious agents were evaluated with the quantitative Luciferase Immunoprecipitation Systems (LIPS) in patients from three disease cohorts including those with pathogenic anti-interferon-γ autoantibodies (IFN-γ AAB), HIV and Sjögren’s syndrome (SjS) to determine if their antibody profiles differed from control subjects.  The IFN-γ AAB patients compared to controls demonstrated statistically higher levels of antibodies against VZV (p=0.0003), EBV (p=0.002), CMV (p=0.003), and C. albicans (p=0.03), but lower antibody levels against poliovirus (p=0.04). Comparison of HIV patients with blood donor controls revealed that the patients had higher levels of antibodies against CMV (p=0.0008), HSV-2 (p=0.0008), EBV (p=0.001), and C. albicans (p=0.01), but showed decreased levels of antibodies against coxsackievirus B4 (p=0.0008), poliovirus (p=0.0005),   and HHV-6B (p=0.002). Lastly, SjS patients had higher levels of anti-EBV antibodies (p=0.03), but lower antibody levels against several enteroviruses including a newly identified picornavirus, HCoSV-A (p=0.004), coxsackievirus B4 (p=0.04), and poliovirus (p=0.02). For the IFN-γ AAB and HIV cohorts, principal component analysis revealed unique antibody clusters that showed the potential to discriminate patients from controls.  The results suggest that antibody profiles against these and likely other common infectious agents may yield insight into the interplay between exposure to infectious agents, dysbiosis, adaptive immunity and disease activity. PMID:24312567

  1. Engineered Antibodies for Monitoring of Polynuclear Aromatic Hydrocarbons

    Energy Technology Data Exchange (ETDEWEB)

    Alexander E. Karu Ph.D; Victoria A. Roberts Ph.D.; Qing X. Li, Ph.D.

    2002-01-17

    This project was undertaken to fill needs in ODE's human and ecosystem health effects research, site remediation, rapid emergency response, and regulatory compliance monitoring programs. Doe has greatly stimulated development and validation of antibody-based, rapid, field-portable detection systems for small hazardous compounds. These range from simple dipsticks, microplate enzyme-linked immunosorbent assays (ELISAs), and hand-held colorimeters, to ultrasensitive microfluidic reactors, fiber-optic sensors and microarrays that can identify multiple analytes from patterns of cross-reactivity. Unfortunately, the technology to produce antibodies with the most desirable properties did not keep pace. Lack of antibodies remains a limiting factor in production and practical use of such devices. The goals of our project were to determine the chemical and structural bases for the antibody-analyte binding interactions using advanced computational chemistry, and to use this information to create useful new binding properties through in vitro genetic engineering and combinatorial library methods.

  2. Replacing antibodies with aptamers in lateral flow immunoassay.

    Science.gov (United States)

    Chen, Ailiang; Yang, Shuming

    2015-09-15

    Aptamers have been identified against various targets as a type of chemical or nucleic acid ligand by systematic evolution of ligands by exponential enrichment (SELEX) with high sensitivity and specificity. Aptamers show remarkable advantages over antibodies due to the nucleic acid nature and target-induced structure-switching properties and are widely used to design various fluorescent, electrochemical, or colorimetric biosensors. However, the practical applications of aptamer-based sensing and diagnostics are still lagging behind those of antibody-based tests. Lateral flow immunoassay (LFIA) represents a well established and appropriate technology among rapid assays because of its low cost and user-friendliness. The antibody-based platform is utilized to detect numerous targets, but it is always hampered by the antibody preparation time, antibody stability, and effect of modification on the antibody. Seeking alternatives to antibodies is an area of active research and is of tremendous importance. Aptamers are receiving increasing attention in lateral flow applications because of a number of important potential performance advantages. We speculate that aptamer-based LFIA may be one of the first platforms for commercial use of aptamer-based diagnosis. This review first gives an introduction to aptamer including the selection process SELEX with its focus on aptamer advantages over antibodies, and then depicts LFIA with its focus on aptamer opportunities in LFIA over antibodies. Furthermore, we summarize the recent advances in the development of aptamer-based lateral flow biosensing assays with the aim to provide a general guide for the design of aptamer-based lateral flow biosensing assays. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Effect of tetracycline on transfer and establishment of the tetracycline-inducible conjugative transposon Tn916 in the guts of gnotobiotic rats

    DEFF Research Database (Denmark)

    Bahl, Martin I.; Sørensen, Søren J.; Hansen, Lars H.

    2004-01-01

    and intestinal segments were monitored by using bacterial biosensors carrying a transcriptional fusion of a tetracycline-regulated promoter and a lacZ reporter gene. Chromosomal locations of Tn916 in transconjugants isolated either from the same animal or from different animals were compared by Southern blot...

  4. Establishing exchange bias below T-N with polycrystalline Ni0.52Co0.48O/Co bilayers

    DEFF Research Database (Denmark)

    Berkowitz, A.E.; Hansen, Mikkel Fougt; Tang, Y.J.

    2005-01-01

    Exchange-coupled bilayers of polycrystalline ferromagnetic (FM) Co on antiferromagnetic (AFM) Ni0.52Co0.48O were investigated with emphasis on the issue of establishing an exchange-bias field, H-E, below the AFM ordering temperature, T-N. It was found that field-cooling the bilayers through T......-N provided very little, if any, increase in H-E over that produced by deposition of the Co at temperatures far below T-N. Further significant aspects of this issue were also examined. The biasing field, H-B, needed to be applied only during the deposition of a small fraction (1 nm) of the FM film below T......-N of the AFM to achieve the maximum H-E; if H-B was reversed at any time during the FM deposition, H-E was determined by the final direction of H-B; the direction of H-E could be reversed after it had been established by applying a reversed field during a post-deposition latent period. These and other findings...

  5. High-Level Heat Resistance of Spores of Bacillus amyloliquefaciens and Bacillus licheniformis Results from the Presence of a spoVA Operon in a Tn1546 Transposon

    NARCIS (Netherlands)

    Berendsen, Erwin M; Koning, Rosella A; Boekhorst, Jos; de Jong, Anne; Kuipers, Oscar P; Wells-Bennik, Marjon H J

    2016-01-01

    Bacterial endospore formers can produce spores that are resistant to many food processing conditions, including heat. Some spores may survive heating processes aimed at production of commercially sterile foods. Recently, it was shown that a spoVA operon, designated spoVA(2mob), present on a Tn1546

  6. Application value of combined measurement of serum sTn, CA242, CA19-9 and CEA in the diagnosis of gastroenterological neoplasm

    International Nuclear Information System (INIS)

    Zhang Wanzhong; Chen Zhizhou; Fan Zhenfu

    2007-01-01

    To determine the application value of four serum tumor markers sTn, CA242, CA 19-9 and CEA in the diagnosis of gastroenterological neoplasm, the serum sTn, CA242, CA19-9 and CEA in 30 normal adult controls and 60 patients with gastroenterological neoplasm were measured by IRMA. The results showed that the serum sTn, CA242, CA19-9 and CEA levels in patients with gastric carcinoma or colorectal carcinoma were much higher than those in control group (P<0.01). The serum CEA, CA19-9 and CA242 levels in patients with colorectal carcinoma were significantly higher than those in patients with gastric carcinoma (P<0.01), but the serum sTn level in the former was markedly lower (P<0.01) than that in the latter. The sensitivity of tumor marker increased with the progress of clinical stages, with a considerably higher sensitivity for stage IV compared with stage I-II (P<0.01). The combined test of four tumor markers could be more sensitive than single test in detecting gastric carcinoma and colorectal carcinoma (P<0.05). Four tumor markers are useful for diagnosing gastroenterological neoplasm, and the combined measurement of 4 tumor markers could increase the sensitivity of detecting gastric carcinoma. (authors)

  7. 77 FR 59573 - Proposed Amendment of Class D and E Airspace; Tri-Cities, TN; Revocation of Class E Airspace; Tri...

    Science.gov (United States)

    2012-09-28

    ... at Hawkins County Airport, Rogersville, TN, and Virginia Highlands Airport, Abington, VA. The Tri... separation of existing Class E airspace surrounding Virginia Highlands Airport, Abingdon, VA, and Hawkins... publication, the FAA reassessed the proposal to show the separation of Hawkins County Airport, and Virginia...

  8. The impact of systems-of-care on pharmacoinvasive management with streptokinase: The subgroup analysis of the TN-STEMI programme

    Directory of Open Access Journals (Sweden)

    Deep Chandh Raja

    2017-09-01

    Conclusion: The implementation of a system-of-care (hub-and-spoke model in the pharmacoinvasive group of the TN-STEMI programme demonstrated shorter lysis-to-angiogram times, better TIMI flow patterns and lower thrombus burden in the post-implementation phase.

  9. A new approach for molecular cloning in cyanobacteria: cloning of an anacystis nidulans met gene using a Tn 907-induced mutant

    NARCIS (Netherlands)

    Tandeau de Marsac, N.; Borrias, W.E.; Kuhlemeijer, C.J.; Castets, A.M.; Arkel, G.A. van; Hondel, C.A.M.J.J. van den

    1982-01-01

    A new strategy for molecular cloning in the cyanobacterium Anacystis nidulans R-2 is described. This strategy involved the use of a transposon and was developed for the cloning of a gene encoding methionine biosynthesis. A met::Tn 901 mutant was isolated. Chromosomal DNA fragments were cloned in the

  10. Immunization with Clinical HIV-1 Env Proteins Induces Broad Antibody Dependent Cellular Cytotoxicity-Mediating Antibodies in a Rabbit Vaccination Model

    DEFF Research Database (Denmark)

    Karlsson, Ingrid; Borggren, Marie; Jensen, Sanne Skov

    2018-01-01

    The induction of both neutralizing antibodies and non-neutralizing antibodies with effector functions, for example, antibody-dependent cellular cytotoxicity (ADCC), is desired in the search for effective vaccines against HIV-1. In the pursuit of novel immunogens capable of inducing an efficient...... of inducing antibodies with ADCC activity did not necessarily induce neutralizing activity and vice versa. Nevertheless, we identified vaccine candidates that were able to concurrently induce both types of responses and that had ADCC activity that was cross-reactive between different subtypes. When searching...

  11. Detección del antígeno Tn en tumores epiteliales con la lectina de Vicia villosa isolectina B4.

    Directory of Open Access Journals (Sweden)

    Catalina Limpias

    2010-10-01

    Full Text Available Antecedentes. Los epítopes T, Tn y sTn, se expresan en un alto porcentaje de tumores epiteliales y pueden detectarse con anticuerpos monoclonales y lectinas. Objetivo. Evaluar diferencias de expresión del antígeno Tn en cortes histológicos de epitelios no neoplásicos y tumores epiteliales mediante isolectina B4 de Vicia villosa. Material y métodos. Se evaluaron semicuantitativamente localización, intensidad y porcentaje de expresión del antígeno en carcinomas in-situ e infiltrantes y epitelios no neoplásicos de cérvix, seno y urotelio, mediante isolectina B4. Resultados La expresión de Tn en cérvix predominó en membrana de células no neoplásicas y citoplasma de células tumorales; su intensidad fue mayor en carcinomas in-situ e infiltrantes comparado con epitelio no neoplásico aunque en este el porcentaje de expresión fue mayor. En seno, la expresión de Tn fue predominantemente citoplasmática con intensidad similar, el porcentaje de expresión fué mayor en carcinomas ductales in-situ e infiltrantes. En urotelio no neoplásico y tumoral la expresión de Tn predominó en citoplasma; la intensidad y el porcentaje de expresión fueron mayores en neoplasias no invasivas de bajo y alto grado, mientras que en urotelio no neoplásico fue baja y no hubo tendencia definida en tumores infiltrantes. Conclusiones. La detección del antígeno Tn mediante la lectina VVB4 mostró una mayor extensión de marcación en carcinomas ductales de seno en relación con el epitelio no neoplásico, pero no mostró una tendencia definida entre el tejido normal, ni diferentes etapas del desarrollo de los tumores de cérvix y urotelio. Estos hallazgos pueden atribuirse a la heterogeneidad de los procesos carcinogénicos o a que la especificidad de la lectina VVB4 no está restringida a este antígeno.

  12. Antibody screening in multitransfused patients: a prerequisite before each transfusion.

    Science.gov (United States)

    Lamba, Divjot S; Mittal, Kshitija; Sood, Tanvi; Bedi, Ravneet Kaur; Kaur, Paramjit; Kaur, Gagandeep

    2014-10-01

    Life-long red blood cell (RBC) transfusions remain the main treatment for severe thalassemia. We hereby report a case of anti S and anti Lu(a) in a β-thalassemia major patient detected incidentally on antibody screening. The patient was a known case of β-thalassemia major and was on regular blood transfusion every 3 weeks from the institute from the age of 6 months. Subsequently, on one occasion, patient's crossmatch was compatible despite positive antibody screen using microcolumn gel technique. Autocontrol and direct antiglobulin test were negative on microcolumn gel. Anti S and anti Lu(a) antibodies were identified. Blood unit found compatible was negative for S and Lu(a) antigens. Antibody titers were 1:1 for both anti S and anti Lu(a) in AHG phase using tube technique and antibodies were of IgG type. Blood unit was transfused uneventfully to the patient. Donors were traced back (last three donations) and called for repeat blood sample testing for S and Lu(a) antigen. Two out of three donors were found to be S antigen positive and one out of these two was Lu(a) antigen positive. Anti S and anti Lu(a) antibodies were again identified on patient's subsequent visit for transfusion. The present case re-emphasize the importance of antibody screening at each visit in earlier detection of antibodies in multi transfused patients. Encouraging patients to receive transfusion from one center and dedicating donors could reduce alloimmunization rate but larger studies are required. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. New tests to detect antiphospholipid antibodies: antiprothrombin (aPT) and anti-phosphatidylserine/prothrombin (aPS/PT) antibodies.

    Science.gov (United States)

    Sciascia, Savino; Khamashta, Munther A; Bertolaccini, Maria Laura

    2014-05-01

    Antiprothrombin antibodies have been proposed as potential new biomarkers for thrombosis and/or pregnancy morbidity in the setting of the antiphospholipid syndrome (APS). Antiprothrombin antibodies are commonly detected by ELISA, using prothrombin coated onto irradiated plates (aPT), or prothrombin in complex with phosphatidylserine (aPS/PT), as antigen. Although these antibodies can co-exist in the same patient, aPT and aPS/PT seem to belong to different populations of autoantibodies. Early research explored the role of antibodies to prothrombin as potential antigenic targets for the lupus anticoagulant (LA). To date their clinical significance is being investigated and their potential role in identifying patients at higher risk of developing thrombotic events or pregnancy morbidity is being probed.

  14. Analysis of 303 Ro/SS-A antibody-positive patients: is this antibody a possible marker for malignancy?

    Science.gov (United States)

    Böckle, B C; Stanarevic, G; Ratzinger, G; Sepp, N T

    2012-11-01

    The risk of cancer in patients with autoimmune diseases has been investigated in several studies. Ro/SS-A antibodies are frequent and specific autoantibodies among patients with various autoimmune diseases. To assess the risk of cancer in individuals with positive Ro/SS-A antibodies and to analyse their clinical and laboratory characteristics. Consecutive patients (n = 303) with Ro/SS-A antibody positivity were collected during 11 years in our outpatient clinic for autoimmune diseases and were retrospectively analysed. Standardized incidence ratios (SIRs) and 95% confidence intervals (CIs) for all cancers were calculated. In addition, we identified further clinical and laboratory characteristics of Ro/SS-A antibody-positive patients indicating the development or existence of a malignancy. Fifty (16·5%) patients were diagnosed with malignancies. Ro/SS-A antibody was strongly associated with malignant diseases (SIR 2·6, 95% CI 1·9-6·1), particularly melanoma (SIR 33·3, 95% CI 5·2-188·6), T-cell lymphoma (SIR 16·7, 95% CI 2·9-128·9), non-Hodgkin lymphoma (SIR 10·6, 95% CI 1·5-78·9) and breast carcinoma (SIR 4·98, 95% CI 1·3-28·3). Logistic regression modelling revealed that Ro/SS-A antibody-positive patients aged 55 years or older, presenting with fever, anaemia and cutaneous lupus erythematosus, have a greater probability of developing cancer and are considered high-risk patients, as compared with Ro/SS-A antibody-positive patients with none of the mentioned clinical criteria. In our cohort of Ro/SS-A antibody-positive patients, an overall increased risk of malignancy was noticed. Regular screening tests including imaging and laboratory values are justified in Ro/SS-A antibody-positive patients who exhibit the mentioned clinical criteria. © 2012 The Authors. BJD © 2012 British Association of Dermatologists.

  15. Inhibition of human immunodeficiency virus (HIV) infection in vitro by anticarbohydrate monoclonal antibodies

    DEFF Research Database (Denmark)

    Hansen, J E; Clausen, H; Nielsen, C

    1990-01-01

    Carbohydrate structures are often involved in the initial adhesion of pathogens to target cells. In the present study, a panel of anticarbohydrate monoclonal antibodies (MAbs) was tested for their ability to inhibit in vitro human immunodeficiency virus infectivity. MAbs against three different N......- and O-linked carbohydrate epitopes (LeY, A1, and sialyl-Tn) were able to block infection by cell-free virus as well as inhibit syncytium formation. Inhibition of virus infectivity was independent of virus strain (HTLVIIIB or patient isolate SSI-002), the cell line used for virus propagation (H9 or MT4......), and the cell type used as the infection target (MT4, PMC, or selected T4 lymphocytes). Inhibition was observed when viruses were preincubated with MAbs but not when cells were preincubated with MAbs before inoculation, and the MAbs were shown to precipitate 125I-labeled gp120. The MAbs therefore define...

  16. Glycine receptor antibodies in PERM and related syndromes: characteristics, clinical features and outcomes

    Science.gov (United States)

    Carvajal-González, Alexander; Leite, M. Isabel; Waters, Patrick; Woodhall, Mark; Coutinho, Ester; Balint, Bettina; Lang, Bethan; Pettingill, Philippa; Carr, Aisling; Sheerin, Una-Marie; Press, Raomand; Lunn, Michael P.; Lim, Ming; Maddison, Paul; Meinck, H.-M.; Vandenberghe, Wim

    2014-01-01

    The clinical associations of glycine receptor antibodies have not yet been described fully. We identified prospectively 52 antibody-positive patients and collated their clinical features, investigations and immunotherapy responses. Serum glycine receptor antibody endpoint titres ranged from 1:20 to 1:60 000. In 11 paired samples, serum levels were higher than (n = 10) or equal to (n = 1) cerebrospinal fluid levels; there was intrathecal synthesis of glycine receptor antibodies in each of the six pairs available for detailed study. Four patients also had high glutamic acid decarboxylase antibodies (>1000 U/ml), and one had high voltage-gated potassium channel-complex antibody (2442 pM). Seven patients with very low titres (antibodies activated complement on glycine receptor-transfected human embryonic kidney cells at room temperature, and caused internalization and lysosomal degradation of the glycine receptors at 37°C. Immunoglobulin G antibodies bound to rodent spinal cord and brainstem co-localizing with monoclonal antibodies to glycine receptor-α1. Ten glycine receptor antibody positive samples were also identified in a retrospective cohort of 56 patients with stiff person syndrome and related syndromes. Glycine receptor antibodies are strongly associated with spinal and brainstem disorders, and the majority of patients have progressive encephalomyelitis with rigidity and myoclonus. The antibodies demonstrate in vitro evidence of pathogenicity and the patients respond well to immunotherapies, contrasting with earlier studies of this syndrome, which indicated a poor prognosis. The presence of glycine receptor antibodies should help to identify a disease that responds to immunotherapies, but these treatments may need to be sustained, relapses can occur and maintenance immunosuppression may be required. PMID:24951641

  17. Value of serum TORCH-specific antibody detection in assessment of neonatal jaundice

    Directory of Open Access Journals (Sweden)

    Guang-Hua Dai

    2016-06-01

    Full Text Available Objective: To study the value of serum TORCH-specific antibody detection in assessment of neonatal jaundice. Methods: A total of 70 cases of children with neonatal jaundice were selected as jaundice group, 70 cases of healthy newborn were the control group, and serum serum TORCH-specific antibody content as well as heart function, liver function, kidney function and nerve function indicators were detected. Results: Serum TOX-IgM, RV-IgM, CMV-IgM and HSV-IgM positive rate and content of jaundice group were significantly higher than those of control group; serum CK-MB, cTnI, AST, ALT, Cys-C, RBP, MBP, S100β and NSE content of TORCH-positive children were significantly higher than those of TORCHnegative children, and BDNF, NT-3, NT-4 and NGF content were significantly lower than those of TORCH-negative children; T1WI signal of pallidum MRI of TORCH-positive children was significantly higher than that of TORCH-negative children. Conclusions: Serum TORCHspecific antibodies significantly increase in children with neonatal jaundice and can assess the degree of bilirubin metabolism disorder and the degree of target organ damage.

  18. RNA-seq and Tn-seq reveal fitness determinants of vancomycin-resistant Enterococcus faecium during growth in human serum.

    Science.gov (United States)

    Zhang, Xinglin; de Maat, Vincent; Guzmán Prieto, Ana M; Prajsnar, Tomasz K; Bayjanov, Jumamurat R; de Been, Mark; Rogers, Malbert R C; Bonten, Marc J M; Mesnage, Stéphane; Willems, Rob J L; van Schaik, Willem

    2017-11-21

    The Gram-positive bacterium Enterococcus faecium is a commensal of the human gastrointestinal tract and a frequent cause of bloodstream infections in hospitalized patients. The mechanisms by which E. faecium can survive and grow in blood during an infection have not yet been characterized. Here, we identify genes that contribute to growth of E. faecium in human serum through transcriptome profiling (RNA-seq) and a high-throughput transposon mutant library sequencing approach (Tn-seq). We first sequenced the genome of E. faecium E745, a vancomycin-resistant clinical isolate, using a combination of short- and long read sequencing, revealing a 2,765,010 nt chromosome and 6 plasmids, with sizes ranging between 9.3 kbp and 223.7 kbp. We then compared the transcriptome of E. faecium E745 during exponential growth in rich medium and in human serum by RNA-seq. This analysis revealed that 27.8% of genes on the E. faecium E745 genome were differentially expressed in these two conditions. A gene cluster with a role in purine biosynthesis was among the most upregulated genes in E. faecium E745 upon growth in serum. The E. faecium E745 transposon mutant library was then used to identify genes that were specifically required for growth of E. faecium in serum. Genes involved in de novo nucleotide biosynthesis (including pyrK_2, pyrF, purD, purH) and a gene encoding a phosphotransferase system subunit (manY_2) were thus identified to be contributing to E. faecium growth in human serum. Transposon mutants in pyrK_2, pyrF, purD, purH and manY_2 were isolated from the library and their impaired growth in human serum was confirmed. In addition, the pyrK_2 and manY_2 mutants were tested for their virulence in an intravenous zebrafish infection model and exhibited significantly attenuated virulence compared to E. faecium E745. Genes involved in carbohydrate metabolism and nucleotide biosynthesis of E. faecium are essential for growth in human serum and contribute to the pathogenesis of

  19. Mucoepidermoid carcinoma-associated expression of MUC5AC, MUC5B and mucin-type carbohydrate antigen sialyl-Tn in the parotid gland.

    Science.gov (United States)

    Matse, Johannes H; Bharos, Wiresh K; Veerman, Enno C I; Bloemena, Elisabeth; Bolscher, Jan G M

    2017-10-01

    The aberrant expression of mucins and mucin-type carbohydrates has been described in many types of cancer, including mucoepidermoid carcinoma (MEC), a malignant salivary gland tumor. In this study, we examined the aberrant expression patterns of mucins (MUC1, MUC4, MUC5AC and MUC5B), simple mucin-type carbohydrate antigens (Tn, sialyl-Tn and T) and mature carbohydrate antigens (Lewis a and sulfo-Lewis a antigens) in MEC originating from the parotid gland, which normally does not secrete mucins. We conducted an immunohistochemical study to investigate the presence of mucins and carbohydrates in 24 MEC samples originating from the parotid gland and in surrounding normal tissue of the same gland in comparison 6 samples of normal salivary glands. The expression levels were compared with respect to the histological grading. Furthermore, 24 MEC samples from non-parotid salivary glands were included. We observed loss of topology of membrane-bound MUC1 and MUC4, and de novo expression of MUC5AC, MUC5B and sialyl-Tn in MEC that originated in the parotid gland. Furthermore, mucins MUC1, MUC4 and carbohydrate antigens Tn, sialyl-Tn, T, Lewis a and sulfo-Lewis a were overexpressed in MEC samples compared to surrounding normal salivary gland tissues. MUC1 was expressed in both low- and high grade MECs, whereas MUC4 was not expressed in high grade MECs of the parotid gland. During the development of MEC in the parotid gland, the genes for gel-forming secretory mucins are switched on. Besides these MEC tissues overexpress short oligosaccharides, suggesting that the glycosylation machinery is altered. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Troponin C Mutations Partially Stabilize the Active State of Regulated Actin and Fully Stabilize the Active State When Paired with Δ14 TnT.

    Science.gov (United States)

    Baxley, Tamatha; Johnson, Dylan; Pinto, Jose R; Chalovich, Joseph M

    2017-06-13

    Striated muscle contraction is regulated by the actin-associated proteins tropomyosin and troponin. The extent of activation of myosin ATPase activity is lowest in the absence of both Ca 2+ and activating cross-bridges (i.e., S1-ADP or rigor S1). Binding of activating species of myosin to actin at a saturating Ca 2+ concentration stabilizes the most active state (M state) of the actin-tropomyosin-troponin complex (regulated actin). Ca 2+ binding alone produces partial stabilization of the active state. The extent of stabilization at a saturating Ca 2+ concentration depends on the isoform of the troponin subunits, the phosphorylation state of troponin, and, in the case of cardiac muscle, the presence of hypertrophic cardiomyopathy-producing mutants of troponin T and troponin I. Cardiac dysfunction is also associated with mutations of troponin C (TnC). Troponin C mutants A8V, C84Y, and D145E increase the Ca 2+ sensitivity of ATPase activity. We show that these mutants change the distribution of regulated actin states. The A8V and C84Y TnC mutants decreased the inactive B state distribution slightly at low Ca 2+ concentrations, but the D145E mutants had no effect on that state. All TnC mutants increased the level of the active M state compared to that of the wild type, at a saturating Ca 2+ concentration. Troponin complexes that contained two mutations that stabilize the active M state, A8V TnC and Δ14 TnT, appeared to be completely in the active state in the presence of only Ca 2+ . Because Ca 2+ gives full activation, in this situation, troponin must be capable of positioning tropomyosin in the active M state without the need for rigor myosin binding.

  1. High-Level Heat Resistance of Spores of Bacillus amyloliquefaciens and Bacillus licheniformis Results from the Presence of a spoVA Operon in a Tn1546 Transposon

    Science.gov (United States)

    Berendsen, Erwin M.; Koning, Rosella A.; Boekhorst, Jos; de Jong, Anne; Kuipers, Oscar P.; Wells-Bennik, Marjon H. J.

    2016-01-01

    Bacterial endospore formers can produce spores that are resistant to many food processing conditions, including heat. Some spores may survive heating processes aimed at production of commercially sterile foods. Recently, it was shown that a spoVA operon, designated spoVA2mob, present on a Tn1546 transposon in Bacillus subtilis, leads to profoundly increased wet heat resistance of B. subtilis spores. Such Tn1546 transposon elements including the spoVA2mob operon were also found in several strains of Bacillus amyloliquefaciens and Bacillus licheniformis, and these strains were shown to produce spores with significantly higher resistances to wet heat than their counterparts lacking this transposon. In this study, the locations and compositions of Tn1546 transposons encompassing the spoVA2mob operons in B. amyloliquefaciens and B. licheniformis were analyzed. Introduction of these spoVA2mob operons into B. subtilis 168 (producing spores that are not highly heat resistant) rendered mutant 168 strains that produced high-level heat resistant spores, demonstrating that these elements in B. amyloliquefaciens and B. licheniformis are responsible for high level heat resistance of spores. Assessment of growth of the nine strains of each species between 5.2°C and 57.7°C showed some differences between strains, especially at lower temperatures, but all strains were able to grow at 57.7°C. Strains of B. amyloliquefaciens and B. licheniformis that contain the Tn1546 elements (and produce high-level heat resistant spores) grew at temperatures similar to those of their Tn1546-negative counterparts that produce low-level heat resistant spores. The findings presented in this study allow for detection of B. amyloliquefaciens and B. licheniformis strains that produce highly heat resistant spores in the food chain. PMID:27994575

  2. Tn5253 family integrative and conjugative elements carrying mef(I) and catQ determinants in Streptococcus pneumoniae and Streptococcus pyogenes.

    Science.gov (United States)

    Mingoia, Marina; Morici, Eleonora; Morroni, Gianluca; Giovanetti, Eleonora; Del Grosso, Maria; Pantosti, Annalisa; Varaldo, Pietro E

    2014-10-01

    The linkage between the macrolide efflux gene mef(I) and the chloramphenicol inactivation gene catQ was first described in Streptococcus pneumoniae (strain Spn529), where the two genes are located in a module designated IQ element. Subsequently, two different defective IQ elements were detected in Streptococcus pyogenes (strains Spy029 and Spy005). The genetic elements carrying the three IQ elements were characterized, and all were found to be Tn5253 family integrative and conjugative elements (ICEs). The ICE from S. pneumoniae (ICESpn529IQ) was sequenced, whereas the ICEs from S. pyogenes (ICESpy029IQ and ICESpy005IQ, the first Tn5253-like ICEs reported in this species) were characterized by PCR mapping, partial sequencing, and restriction analysis. ICESpn529IQ and ICESpy029IQ were found to share the intSp 23FST81 integrase gene and an identical Tn916 fragment, whereas ICESpy005IQ has int5252 and lacks Tn916. All three ICEs were found to lack the linearized pC194 plasmid that is usually associated with Tn5253-like ICEs, and all displayed a single copy of a toxin-antitoxin operon that is typically contained in the direct repeats flanking the excisable pC194 region when this region is present. Two different insertion sites of the IQ elements were detected, one in ICESpn529IQ and ICESpy029IQ, and another in ICESpy005IQ. The chromosomal integration of the three ICEs was site specific, depending on the integrase (intSp 23FST81 or int5252). Only ICESpy005IQ was excised in circular form and transferred by conjugation. By transformation, mef(I) and catQ were cotransferred at a high frequency from S. pyogenes Spy005 and at very low frequencies from S. pneumoniae Spn529 and S. pyogenes Spy029. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  3. High-level heat resistance of spores of Bacillus amyloliquefaciens and Bacillus licheniformis results from the presence of a spoVA operon in a Tn1546 transposon.

    Directory of Open Access Journals (Sweden)

    Erwin M. Berendsen

    2016-12-01

    Full Text Available Bacterial endospore formers can produce spores that are resistant to many food processing conditions, including heat. Some spores may survive heating processes aimed at production of commercially sterile foods. Recently, it was shown that a spoVA operon, designated spoVA2mob, present on a Tn1546 transposon in Bacillus subtilis, leads to profoundly increased wet heat resistance of B. subtilis spores. Such Tn1546 transposon elements including the spoVA2mob operon were also found in several strains of Bacillus amyloliquefaciens and Bacillus licheniformis, and these strains were shown to produce spores with significantly higher resistances to wet heat than their counterparts lacking this transposon. In this study, the locations and compositions of Tn1546 transposons encompassing the spoVA2mob operons in B. amyloliquefaciens and B. licheniformis were analyzed. Introduction of these spoVA2mob operons into B. subtilis 168 (producing spores that are not highly heat resistant rendered mutant 168 strains that produced high-level heat resistant spores, demonstrating that these elements in B. amyloliquefaciens and B. licheniformis are responsible for high level heat resistance of spores. Assessment of growth of the nine strains of each species between 5.2°C and 57.7°C showed some differences between strains, especially at lower temperatures, but all strains were able to grow at 57.7°C. Strains of B. amyloliquefaciens and B. licheniformis that contain the Tn1546 elements (and produce high-level heat resistant spores grew at temperatures similar to those of their Tn1546-negative counterparts that produce low-level heat resistant spores. The findings presented in this study allow for detection of B. amyloliquefaciens and B. licheniformis strains that produce highly heat resistant spores in the food chain.

  4. Antineutrophil Cytoplasmic Antibodies, Autoimmune Neutropenia, and Vasculitis

    Science.gov (United States)

    Grayson, Peter C.; Sloan, J. Mark; Niles, John L.; Monach, Paul A.; Merkel, Peter A.

    2011-01-01

    Objectives Reports of an association between antineutrophil cytoplasmic antibodies (ANCA) and autoimmune neutropenia have rarely included cases of proven vasculitis. A case of ANCA-associated vasculitis (AAV) with recurrent neutropenia is described and relevant literature on the association between ANCA, neutropenia, and vasculitis is reviewed. Methods Longitudinal clinical assessments and laboratory findings are described in a patient with AAV and recurrent episodes of profound neutropenia from December 2008 – October 2010. A PubMed database search of the medical literature was performed for papers published from 1960 through October 2010 to identify all reported cases of ANCA and neutropenia. Results A 49 year-old man developed recurrent neutropenia, periodic fevers, arthritis, biopsy-proven cutaneous vasculitis, sensorineural hearing loss, epididymitis, and positive tests for ANCA with specificity for antibodies to both proteinase 3 and myeloperoxidase. Antineutrophil membrane antibodies were detected during an acute neutropenic phase and were not detectable in a post-recovery sample, whereas ANCA titers did not seem to correlate with neutropenia. An association between ANCA and neutropenia has been reported in 74 cases from 24 studies in the context of drug/toxin exposure, underlying autoimmune disease, or chronic neutropenia without underlying autoimmune disease. In these cases, the presence of atypical ANCA patterns and other antibodies were common; however, vasculitis was uncommon and when it occurred was usually limited to the skin and in cases of underlying toxin exposure. Conclusions ANCA is associated with autoimmune neutropenia, but systemic vasculitis rarely occurs in association with ANCA and neutropenia. The interaction between neutrophils and ANCA may provide insight into understanding both autoimmune neutropenia and AAV. PMID:21507463

  5. Dissecting Immunogenicity of Monoclonal Antibodies

    National Research Council Canada - National Science Library

    Snyder, Christopher

    2002-01-01

    The potential of monoclonal antibodies, (mAbs), for use in therapeutic and diagnostic applications has not been fully realized in part due to counter-immune responses that often arise in patient recipients of mAb...

  6. Dissecting Immunogenicity of Monoclonal Antibodies

    National Research Council Canada - National Science Library

    Snyder, Christopher

    2003-01-01

    The potential of monoclonal antibodies, (mAbs), for use in therapeutic and diagnostic applications has not been fully realized in part due to counter-immune responses that often arise in patient recipients of mAb...

  7. Antisperm antibodies and fertility association.

    Science.gov (United States)

    Restrepo, B; Cardona-Maya, W

    2013-10-01

    To evaluate the relation between antisperm antibodies (ASA) and human fertility by reviewing the scientific literature of the last 45 years. We carried out a review of scientific literature about antisperm antibodies and infertility published in spanish or english in databases as Pubmed, Medline, Scielo, some books and another gray literature include information related to this review and that is published in the last 45 years. Infertile couples suffer infertility by immunological mechanisms mainly by the presence of antisperm antibodies ASA in blood, semen or cervicovaginal secretions; the formation of ASA in men and women may be associated with disturbance in immunomodulatory mechanisms that result in functional impairment of sperm and thus its inability to fertilize the oocyte. Immunological infertility caused by ASA is the result of interference of these antibodies in various stages of fertilization process, inhibiting the ability of interaction between sperm and oocyte. Copyright © 2012 AEU. Published by Elsevier Espana. All rights reserved.

  8. Antibody Drug Conjugates: Preclinical Considerations.

    Science.gov (United States)

    Bornstein, Gadi G

    2015-05-01

    The development path for antibody drug conjugates (ADCs) is more complex and challenging than for unmodified antibodies. While many of the preclinical considerations for both unmodified and antibody drug conjugates are shared, special considerations must be taken into account when developing an ADC. Unlike unmodified antibodies, an ADC must preferentially bind to tumor cells, internalize, and traffic to the appropriate intracellular compartment to release the payload. Parameters that can impact the pharmacological properties of this class of therapeutics include the selection of the payload, the type of linker, and the methodology for payload drug conjugation. Despite a plethora of in vitro assays and in vivo models to screen and evaluate ADCs, the challenge remains to develop improved preclinical tools that will be more predictive of clinical outcome. This review will focus on preclinical considerations for clinically validated small molecule ADCs. In addition, the lessons learned from Mylotarg®, the first in class FDA-approved ADC, are highlighted.

  9. Radiolabeled monoclonal antibodies: a review

    International Nuclear Information System (INIS)

    Toledo e Souza, I.T. de; Okada, H.

    1990-05-01

    Since the description by Kohler and Milstein 1975 of their technique for producing monoclonal antibodies of predefined specificity, it has become a mainstay in most laboratories that utilize immunochemical techniques to study problems in basic, applied or clinical research. Paradoxically, the very success of monoclonal antibodies has generated a literature which is now so vast and scattered that it has become difficult to obtain a perspective. This brief review represents the distillation of many publications relating to the production and use of monoclonaal antibodies as radiopharmaceuticals. Significant advances were made possible in the last few years by combined developments in the fields of tumor-associated antigens and of monoclonal antibodies. In fact monoclonal antibodies against some well defined tumor-associated antigens, has led to significantly greater practical possibilities for producing highly specific radiolabeled antibodies as radiopharmaceuticals for diagnosis and therapy of human tumors. One of the main requirements of this methodology is the availability of stable radiopharmaceutical reagents which after labeling in vivo injection retain the capacity of specific interaction with the defined antigen and their molecular integrity. Since injection into human is the objetive of this kind of study all the specifications of radiopharmaceutical have to be fulfilled e.g. sterility, apirogenicity and absence of toxicity. (author) [pt

  10. Avian Diagnostic and Therapeutic Antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Bradley, David Sherman [UND SMHS

    2012-12-31

    A number of infectious agents have the potential of causing significant clinical symptomology and even death, but dispite this, the number of incidence remain below the level that supports producing a vaccine. Therapeutic antibodies provide a viable treatment option for many of these diseases. We proposed that antibodies derived from West Nile Virus (WNV) immunized geese would be able to treat WNV infection in mammals and potential humans. We demonstrated that WNV specific goose antibodies are indeed successful in treating WNV infection both prophylactically and therapeutically in a golden hamster model. We demonstrated that the goose derived antibodies are non-reactogenic, i.e. do not cause an inflammatory response with multiple exposures in mammals. We also developed both a specific pathogen free facility to house the geese during the antibody production phase and a patent-pending purification process to purify the antibodies to greater than 99% purity. Therefore, the success of these study will allow a cost effective rapidly producible therapeutic toward clinical testing with the necessary infrastructure and processes developed and in place.

  11. Tabhu: tools for antibody humanization.

    KAUST Repository

    Olimpieri, Pier Paolo

    2014-10-09

    SUMMARY: Antibodies are rapidly becoming essential tools in the clinical practice, given their ability to recognize their cognate antigens with high specificity and affinity, and a high yield at reasonable costs in model animals. Unfortunately, when administered to human patients, xenogeneic antibodies can elicit unwanted and dangerous immunogenic responses. Antibody humanization methods are designed to produce molecules with a better safety profile still maintaining their ability to bind the antigen. This can be accomplished by grafting the non-human regions determining the antigen specificity into a suitable human template. Unfortunately, this procedure may results in a partial or complete loss of affinity of the grafted molecule that can be restored by back-mutating some of the residues of human origin to the corresponding murine ones. This trial-and-error procedure is hard and involves expensive and time-consuming experiments. Here we present tools for antibody humanization (Tabhu) a web server for antibody humanization. Tabhu includes tools for human template selection, grafting, back-mutation evaluation, antibody modelling and structural analysis, helping the user in all the critical steps of the humanization experiment protocol. AVAILABILITY: http://www.biocomputing.it/tabhu CONTACT: anna.tramontano@uniroma1.it, pierpaolo.olimpieri@uniroma1.it SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

  12. Radiolabeling and biotinylation of internalizing monoclonal antibody chimeric BR96: Potential use of extracorporeal immunoadsorption with enhanced tumor radioactivity retention of Iodine, Indium and Rhenium

    Energy Technology Data Exchange (ETDEWEB)

    Chen, JianQing

    1997-01-01

    In this thesis, methodology of radiolabeling and simultaneous biotinylation for internalizing monoclonal antibody chimeric BR96 have been investigated by using three element groups of potential therapeutic radionuclides iodine, indium and rhenium, and their different labeling methods. The biodistribution and kinetics of biotinylated and radiolabeled chiBR96 have been studied in colon carcinoma isografted rats. The potential use of ECIA, based on the biotin-avidin concept, has been evaluated and compared with the approach of avidin `chase` in the same animal tumor model with respect to an enhancement of tumor-to-normal tissue (T/N) activity ratio. 131 refs.

  13. Radiolabeling and biotinylation of internalizing monoclonal antibody chimeric BR96: Potential use of extracorporeal immunoadsorption with enhanced tumor radioactivity retention of Iodine, Indium and Rhenium

    International Nuclear Information System (INIS)

    Chen, JianQing.

    1997-01-01

    In this thesis, methodology of radiolabeling and simultaneous biotinylation for internalizing monoclonal antibody chimeric BR96 have been investigated by using three element groups of potential therapeutic radionuclides iodine, indium and rhenium, and their different labeling methods. The biodistribution and kinetics of biotinylated and radiolabeled chiBR96 have been studied in colon carcinoma isografted rats. The potential use of ECIA, based on the biotin-avidin concept, has been evaluated and compared with the approach of avidin 'chase' in the same animal tumor model with respect to an enhancement of tumor-to-normal tissue (T/N) activity ratio. 131 refs

  14. Data Sharing Report Characterization of Isotope Row Facilities Oak Ridge National Laboratory Oak Ridge TN

    Energy Technology Data Exchange (ETDEWEB)

    Weaver, Phyllis C. [Oak Ridge Inst. for Science and Education (ORISE), Oak Ridge, TN (United States)

    2013-12-12

    The U.S. Department of Energy (DOE) Oak Ridge Office of Environmental Management (EM-OR) requested that Oak Ridge Associated Universities (ORAU), working under the Oak Ridge Institute for Science and Education (ORISE) contract, provide technical and independent waste management planning support using funds provided by the American Recovery and Reinvestment Act (ARRA). Specifically, DOE EM-OR requested ORAU to plan and implement a survey approach, focused on characterizing the Isotope Row Facilities located at the Oak Ridge National Laboratory (ORNL) for future determination of an appropriate disposition pathway for building debris and systems, should the buildings be demolished. The characterization effort was designed to identify and quantify radiological and chemical contamination associated with building structures and process systems. The Isotope Row Facilities discussed in this report include Bldgs. 3030, 3031, 3032, 3033, 3033A, 3034, 3036, 3093, and 3118, and are located in the northeast quadrant of the main ORNL campus area, between Hillside and Central Avenues. Construction of the isotope production facilities was initiated in the late 1940s, with the exception of Bldgs. 3033A and 3118, which were enclosed in the early 1960s. The Isotope Row facilities were intended for the purpose of light industrial use for the processing, assemblage, and storage of radionuclides used for a variety of applications (ORNL 1952 and ORAU 2013). The Isotope Row Facilities provided laboratory and support services as part of the Isotopes Production and Distribution Program until 1989 when DOE mandated their shutdown (ORNL 1990). These facilities performed diverse research and developmental experiments in support of isotopes production. As a result of the many years of operations, various projects, and final cessation of operations, production was followed by inclusion into the surveillance and maintenance (S&M) project for eventual decontamination and decommissioning (D&D). The

  15. Waste Disposition Issues and Resolutions at the TRU Waste Processing Center at Oak Ridge TN

    International Nuclear Information System (INIS)

    Gentry, R.

    2009-01-01

    This paper prepared for the Waste Management Conference 2009 provides lessons learned from the Transuranic (TRU) Waste Processing Center (TWPC) associated with development of approaches used to certify and ensure disposition of problematic TRU wastes at the Waste Isolation Pilot Plant (WIPP) site. The TWPC is currently processing the inventory of available waste TRU waste at the Oak Ridge National Lab (ORNL). During the processing effort several waste characteristics were identified/discovered that did not conform to the normal standards and processes for disposal at WIPP. Therefore, the TWPC and ORNL were challenged with determining a path forward for this problematic, special case TRU wastes to ensure that they can be processed, packaged, and shipped to WIPP. Additionally, unexpected specific waste characteristics have challenged the project to identify and develop processing methods to handle problematic waste. The TWPC has several issues that have challenged the projects ability to process RH Waste. High Neutron Dose Rate resulting from both Californium and Curium in the waste stream challenge the RH-TRU 72-B limit for dose rate measured from the side of the package under normal conditions of transport, as specified in Chapter 5.0 of the RH-TRU 72-B SAR (i.e., ≤10 mrem/hour at 2 meters). Difficult to process waste in the hot cell has introduced processing and handling difficulties included problems associated with the disposition of prohibited items that fall out of the waste stream such as liquids, aerosol cans, etc. Lastly, multiple waste streams require characterization and AK challenge the ability to generate dose-to curie models for the waste. Repackaging is one solution to the high neutron dose rate issue. In parallel, an effort is underway to request a change to the TRAMPAC requirements to allow shielding in the drum or canister to reduce the impact of the high neutron dose rates. Due diligence on supporting AK efforts is important in ensuring adequate

  16. Presence of Autoimmune Antibody in Chikungunya Infection

    Directory of Open Access Journals (Sweden)

    Wirach Maek-a-nantawat

    2009-01-01

    Full Text Available Chikungunya infection has recently re-emerged as an important arthropod-borne disease in Thailand. Recently, Southern Thailand was identified as a potentially endemic area for the chikungunya virus. Here, we report a case of severe musculoskeletal complication, presenting with muscle weakness and swelling of the limbs. During the investigation to exclude autoimmune muscular inflammation, high titers of antinuclear antibody were detected. This is the report of autoimmunity detection associated with an arbovirus infection. The symptoms can mimic autoimmune polymyositis disease, and the condition requires close monitoring before deciding to embark upon prolonged specific treatment with immunomodulators.

  17. Utility of HLA Antibody Testing in Kidney Transplantation

    Science.gov (United States)

    Konvalinka, Ana

    2015-01-01

    HLA antigens are polymorphic proteins expressed on donor kidney allograft endothelium and are critical targets for recipient immune recognition. HLA antibodies are risk factors for acute and chronic rejection and allograft loss. Solid-phase immunoassays for HLA antibody detection represent a major advance in sensitivity and specificity over cell-based methods and are widely used in organ allocation and pretransplant risk assessment. Post-transplant, development of de novo donor–specific HLA antibodies and/or increase in donor-specific antibodies from pretransplant levels are associated with adverse outcomes. Although single antigen bead assays have allowed sensitive detection of recipient HLA antibodies and their specificities, a number of interpretive considerations must be appreciated to understand test results in clinical and research contexts. This review, which is especially relevant for clinicians caring for transplant patients, discusses the technical aspects of single antigen bead assays, emphasizes their quantitative limitations, and explores the utility of HLA antibody testing in identifying and managing important pre- and post-transplant clinical outcomes. PMID:25804279

  18. Monoclonal antibodies to drosophila cytochrome P-450's

    Energy Technology Data Exchange (ETDEWEB)

    Sundseth, S.S.; Kennel, S.J.; Waters, L.C.

    1987-05-01

    Hybridomas producing monoclonal antibodies were prepared by the fusion of SP2/0 myeloma cells and spleen cells from a female BALB/c mouse immunized by cytochrome P-450-A and P-450-B purified from Drosophila Hikone-R (BG) microsomes. P-450-A and P-450-B are electrophoretically distinct subsets of Drosophila P-450. P-450-A is ubiquitous among strains tested, while P-450-B is present in only a few strains displaying unique enzyme activities and increased insecticide resistance. The Oregon-R strain contains only cytochromes P-450-A and is susceptible to insecticides. The authors Hikone-R (BG) strain expresses both cytochromes P-450-A and P-450-B and is insecticide resistant. Antibody producing hybridomas were detected in a solid-phase radioimmunoassay (RIA) by binding to Hikone-R (BG) or Oregon-R microsomes. Four independent hybridomas were identified as producing monoclonal antibodies that recognized proteins in the P-450 complex by immunoblot experiments. Three monoclonal antibodies recognized P-450-A proteins, while one monoclonal antibody bound predominantly P-450-B. This monoclonal antibody also recognized southern armyworm (Spodoptera eridania, Cramer) microsomal proteins.

  19. Development of Broadly Neutralizing Antibody Mimitopes for Characterization of CRF01_AE HIV-1 Antibody Responses

    Directory of Open Access Journals (Sweden)

    Jesse V. Schoen

    2017-10-01

    Full Text Available Mapping humoral immune responses to HIV-1 over the course of natural infection is important in understanding epitope exposure in relation to elicitation of broadly neutralizing antibodies (bNAbs, which is considered imperative for effective vaccine design. When analyzing HIV-specific immune responses, the antibody binding profiles may be a correlate for functional antibody activity. In this study, we utilized phage display technology to identify novel mimitopes that may represent Env epitope structures bound by bNAbs directed at V1V2 and V3 domains, CD4 binding site (CD4bs and the membrane proximal external region (MPER of Env. Mimitope sequence motifs were determined for each bNAb epitope. Given the ongoing vaccine development efforts in Thailand, these mimitopes that represent CD4bs and MPER epitopes were used to map immune responses of HIV-1 CRF01_AE-infected individuals with known neutralizing responses from two distinct time periods, 1996-98 and 2012-15. The more contemporary cohort showed an increase in binding breadth with binding observed for all MPER and CD4bs mimitopes, while the older cohort showed only 75% recognition of the CD4bs mimitopes and no MPER mimotope binding. Furthermore, mimitope binding profiles correlated significantly with magnitude (p=0.0036 and breadth (p=0.0358 of neutralization of a multi-subtype Tier 1 panel of pseudoviruses. These results highlight the utility of this mimitope mapping approach for detecting human plasma IgG-specificities that target known neutralizing antibody epitopes, and may also provide an indication of the plasticity of antibody binding within HIV-1 Env neutralization determinants.

  20. GABARAPL1 antibodies: target one protein, get one free!

    Science.gov (United States)

    Le Grand, Jaclyn Nicole; Chakrama, Fatima Zahra; Seguin-Py, Stéphanie; Fraichard, Annick; Delage-Mourroux, Régis; Jouvenot, Michèle; Risold, Pierre-Yves; Boyer-Guittaut, Michaël

    2011-11-01

    Atg8 is a yeast protein involved in the autophagic process and in particular in the elongation of autophagosomes. In mammals, several orthologs have been identified and are classed into two subfamilies: the LC3 subfamily and the GABARAP subfamily, referred to simply as the LC3 or GABARAP families. GABARAPL1 (GABARAP-like protein 1), one of the proteins belonging to the GABARAP (GABA(A) receptor-associated protein) family, is highly expressed in the central nervous system and implicated in processes such as receptor and vesicle transport as well as autophagy. The proteins that make up the GABARAP family demonstrate conservation of their amino acid sequences and protein structures. In humans, GABARAPL1 shares 86% identity with GABARAP and 61% with GABARAPL2 (GATE-16). The identification of the individual proteins is thus very limited when working in vivo due to a lack of unique peptide sequences from which specific antibodies can be developed. Actually, and to our knowledge, there are no available antibodies on the market that are entirely specific to GABARAPL1 and the same may be true of the anti-GABARAP antibodies. In this study, we sought to examine the specificity of three antibodies targeted against different peptide sequences within GABARAPL1: CHEM-CENT (an antibody raised against a short peptide sequence within the center of the protein), PTG-NTER (an antibody raised against the N-terminus of the protein) and PTG-FL (an antibody raised against the full-length protein). The results described in this article demonstrate the importance of testing antibody specificity under the conditions for which it will be used experimentally, a caution that should be taken when studying the expression of the GABARAP family proteins.

  1. Genetically engineered multivalent single chain antibody constructs for cancer therapy

    Energy Technology Data Exchange (ETDEWEB)

    Surinder Batra, Ph D

    2006-02-27

    Current therapeutic approaches against the advanced stages of human solid tumors are palliative rather than curative. Many modalities, including, surgery, radiation, and chemotherapy, either alone or in combination have met with only modest success for advanced metastatic cancers. Radioimmunotherapy (RIT) combines the specificity of monoclonal antibodies with cytotxic effects of radioisotopes. It is the smart way of delivering radiation to the known and occult metastatic cancer cells and is independent of drug toxicity and/or hormone resistance. The tumor associated glycoprotein-72 (TAG-72) containing the unique disaccharide sialyl-Tn, is highly expressed in majority of adenocarcinomas, including carcinomas of the prostate, breast, ovaries, pancreas and colon (80-90%) compared to undetectable expression in normal tissues. Monoclonal antibody CC49, reactive with TAG-72, after conjugation to potent gamma- and beta-emitting radionuclides, has been useful in selective systemic radiolocalization of disease and therapy of primary and metastatic tumor sites. However, limited therapeutic responses were observed in patients. Limited success of antibody based delivery of radioisotopes can be attributed to several factors including undesirable pharmacokinetics, poor tumor uptake and high immunogenicity of intact antibodies (IgGs). The primary factors contributing towards the failure of RIT include: 1) longer serum half-lives of the intact IgG molecules resulting in the radiotoxicity, 2) generation of human antibodies against murine antibodies (HAMA) that limits the frequency of dose administration, 3) poor diffusion rates of intact IgG due to the large size and 4) high interstitial fluid pressures (IFP) encountered in solid tumors. The major goal of our multidisciplinary project was to develop specific novel radiopharmaceuticals, with desired pharmacokinetics, for the diagnosis and therapy of solid tumors. To overcome the low uptake of radioactivity by tumors and to increase

  2. Genetically engineered multivalent single chain antibody constructs for cancer therapy

    International Nuclear Information System (INIS)

    Surinder Batra

    2006-01-01

    Current therapeutic approaches against the advanced stages of human solid tumors are palliative rather than curative. Many modalities, including, surgery, radiation, and chemotherapy, either alone or in combination have met with only modest success for advanced metastatic cancers. Radioimmunotherapy (RIT) combines the specificity of monoclonal antibodies with cytotoxic effects of radioisotopes. It is the ''smart'' way of delivering radiation to the known and occult metastatic cancer cells and is independent of drug toxicity and/or hormone resistance. The tumor associated glycoprotein-72 (TAG-72) containing the unique disaccharide sialyl-Tn, is highly expressed in majority of adenocarcinomas, including carcinomas of the prostate, breast, ovaries, pancreas and colon (80-90%) compared to undetectable expression in normal tissues. Monoclonal antibody CC49, reactive with TAG-72, after conjugation to potent gamma- and beta-emitting radionuclides, has been useful in selective systemic radiolocalization of disease and therapy of primary and metastatic tumor sites. However, limited therapeutic responses were observed in patients. Limited success of antibody based delivery of radioisotopes can be attributed to several factors including undesirable pharmacokinetics, poor tumor uptake and high immunogenicity of intact antibodies (IgGs). The primary factors contributing towards the failure of RIT include: (1) longer serum half-lives of the intact IgG molecules resulting in the radiotoxicity, (2) generation of human antibodies against murine antibodies (HAMA) that limits the frequency of dose administration, (3) poor diffusion rates of intact IgG due to the large size and (4) high interstitial fluid pressures (IFP) encountered in solid tumors. The major goal of our multidisciplinary project was to develop specific novel radiopharmaceuticals, with desired pharmacokinetics, for the diagnosis and therapy of solid tumors. To overcome the low uptake of radioactivity by tumors and to

  3. Development of stable reporter system cloning luxCDABE genes into chromosome of Salmonella enterica serotypes using Tn7 transposon

    Directory of Open Access Journals (Sweden)

    Lawrence Mark L

    2010-07-01

    Full Text Available Abstract Background Salmonellosis may be a food safety problem when raw food products are mishandled and not fully cooked. In previous work, we developed bioluminescent Salmonella enterica serotypes using a plasmid-based reporting system that can be used for real-time monitoring of the pathogen's growth on food products in short term studies. In this study, we report the use of a Tn7-based transposon system for subcloning of luxCDABE genes into the chromosome of eleven Salmonella enterica serotypes isolated from the broiler production continuum. Results We found that the lux operon is constitutively expressed from the chromosome post-transposition and the lux cassette is stable without external pressure, i.e. antibiotic selection, for all Salmonella enterica serotypes used. Bioluminescence expression is based on an active electron transport chain and is directly related with metabolic activity. This relationship was quantified by measuring bioluminescence against a temperature gradient in aqueous solution using a luminometer. In addition, bioluminescent monitoring of two serotypes confirmed that our chicken skin model has the potential to be used to evaluate pathogen mitigation strategies. Conclusions This study demonstrated that our new stable reporting system eliminates bioluminescence variation due to plasmid instability and provides a reliable real-time experimental system to study application of preventive measures for Salmonella on food products in real-time for both short and long term studies.

  4. Development of a new neutron shielding material, TN trademark Resin Vyal for transport/storage casks for radioactive materials

    International Nuclear Information System (INIS)

    Abadie, P.

    2004-01-01

    TN trademark Resin Vyal, a patent pending composite, is a new neutron shielding material developed by COGEMA LOGISTICS for transport/storage casks of radioactive materials (spent fuel, reprocessed fuel,..). This material is composed of a thermosetting resin (vinylester resin in solution of styrene) and two mineral fillers (alumine hydrate and zinc borate). Its shielding ability for neutron radiation is related to a high hydrogen content (for slowing down neutrons) and a high boron content (for absorbing neutrons). Source of hydrogen is organic matrix (resin) and alumine hydrate; source of boron is zinc borate. Atomic concentrations are equal to 5.10 22 at/cm 3 for hydrogen and 9.10 20 at/cm 3 for boron. Due to the flame retardant character of components, the final material has a good fire resistance (it is auto-extinguishable). Its density is equal to 1,8. The manufacturing process of these material is easy: it consists in mixing the fillers and pouring in-situ (in cask); so, the curing of this composite, which leads to a three-dimensional structure, takes place at ambient temperature. Temperature resistance of this material was evaluated by performing exposition tests of samples at different temperatures (150 C to 170 C). According to tests results, its maximal temperature of use was confirmed equal to 160 C

  5. Radioiodination of antibodies for tumor imaging

    International Nuclear Information System (INIS)

    Saha, G.B.

    1983-01-01

    In view of the great potential of radioiodinated antibody for the detection and treatment of cancer, the present article deals with the various techniques of radioiodination of antibody and their uses. Topics include methods of iodination of antibody, advantages and disadvantages of different methods, and effects of radioiodination on the antibody molecules with respect to their physiochemical and immunologic reactivity. In addition, the clinical usefulness of radioiodinated antibodies is discussed. (Auth.)

  6. Antibodies from plants for bionanomaterials.

    Science.gov (United States)

    Edgue, Gueven; Twyman, Richard M; Beiss, Veronique; Fischer, Rainer; Sack, Markus

    2017-11-01

    Antibodies are produced as part of the vertebrate adaptive immune response and are not naturally made by plants. However, antibody DNA sequences can be introduced into plants, and together with laboratory technologies that allow the design of antibodies recognizing any conceivable molecular structure, plants can be used as 'green factories' to produce any antibody at all. The advent of plant-based transient expression systems in particular allows the rapid, convenient, and safe production of antibodies, ranging from laboratory-scale expression to industrial-scale manufacturing. The key features of plant-based production include safety, speed, low cost, and convenience, allowing newcomers to rapidly master the technology and use it to its full advantage. Manufacturing in plants has recently achieved significant milestones and offers more than just an alternative to established microbial and mammalian cell platforms. The use of plants for product development in particular offers the power and flexibility to easily coexpress many different genes, allowing the plug-and-play construction of novel bionanomaterials, perfectly complementing existing approaches based on plant virus-like particles. As well as producing single antibodies for applications in medicine, agriculture, and industry, plants can be used to produce antibody-based supramolecular structures and scaffolds as a new generation of green bionanomaterials that promise a bright future based on clean and renewable nanotechnology applications. WIREs Nanomed Nanobiotechnol 2017, 9:e1462. doi: 10.1002/wnan.1462 For further resources related to this article, please visit the WIREs website. © 2017 The Authors. WIREs Nanomedicine and Nanobiotechnology published by Wiley Periodicals, Inc.

  7. Antibody-Directed Phototherapy (ADP

    Directory of Open Access Journals (Sweden)

    M. Adil Butt

    2013-04-01

    Full Text Available Photodynamic therapy (PDT is a clinically-approved but rather under-exploited treatment modality for cancer and pre-cancerous superficial lesions. It utilises a cold laser or LED to activate a photochemical reaction between a light activated drug (photosensitiser-drug and oxygen to generate cytotoxic oxygen species. These free radical species damage cellular components leading to cell death. Despite its benefits, the complexity, limited potency and side effects of PDT have led to poor general usage. However, the research area is very active with an increasing understanding of PDT-related cell biology, photophysics and significant progress in molecular targeting of disease. Monoclonal antibody therapy is maturing and the next wave of antibody therapies includes antibody-drug conjugates (ADCs, which promise to be more potent and curable. These developments could lift antibody-directed phototherapy (ADP to success. ADP promises to increase specificity and potency and improve drug pharmacokinetics, thus delivering better PDT drugs whilst retaining its other benefits. Whole antibody conjugates with first generation ADP-drugs displayed problems with aggregation, poor pharmacokinetics and loss of immuno-reactivity. However, these early ADP-drugs still showed improved selectivity and potency. Improved PS-drug chemistry and a variety of conjugation strategies have led to improved ADP-drugs with retained antibody and PS-drug function. More recently, recombinant antibody fragments have been used to deliver ADP-drugs with superior drug loading, more favourable pharmacokinetics, enhanced potency and target cell selectivity. These improvements offer a promise of better quality PDT drugs.

  8. Antibody Validation by Western Blotting.

    Science.gov (United States)

    Signore, Michele; Manganelli, Valeria; Hodge, Alex

    2017-01-01

    Validation of antibodies is an integral part of translational research, particularly for biomarker discovery. Assaying the specificity of the reagent (antibody) and confirming the identity of the protein biomarker is of critical importance prior to implementing any biomarker in clinical studies, and the lack of such quality control tests may result in unexpected and/or misleading results.Antibody validation is the procedure in which a single antibody is thoroughly assayed for sensitivity and specificity. Although a plethora of commercial antibodies exist, antibody specificity must be extensively demonstrated using diverse complex biological samples, rather than purified recombinant proteins, prior to use in clinical translational research. In the simplest iteration, antibody specificity is determined by the presence of a single band in a complex biological sample, at the expected molecular weight, on a Western blot.To date, numerous Western blotting procedures are available, based on either manual or automated systems and spanning the spectrum of single blots to multiplex blots. X-ray film is still employed in many research laboratories, but digital imaging has become a gold standard in immunoblotting. The basic principles of Western blotting are (a) separation of protein mixtures by gel electrophoresis, (b) transfer of the proteins to a blot, (c) probing the blot for a protein or proteins of interest, and (d) subsequent detection of the protein by chemiluminescent, fluorescent, or colorimetric methods. This chapter focuses on the chemiluminescent detection of proteins using a manual Western blotting system and a vacuum-enhanced detection system (SNAP i.d.™, Millipore).

  9. Staphylococcus aureus virulence factors identified by using a high-throughput Caenorhabditis elegans-killing model.

    Science.gov (United States)

    Begun, Jakob; Sifri, Costi D; Goldman, Samuel; Calderwood, Stephen B; Ausubel, Frederick M

    2005-02-01

    Staphylococcus aureus is an important human pathogen that is also able to kill the model nematode Caenorhabditis elegans. We constructed a 2,950-member Tn917 transposon insertion library in S. aureus strain NCTC 8325. Twenty-one of these insertions exhibited attenuated C. elegans killing, and of these, 12 contained insertions in different genes or chromosomal locations. Ten of these 12 insertions showed attenuated killing phenotypes when transduced into two different S. aureus strains, and 5 of the 10 mutants correspond to genes that have not been previously identified in signature-tagged mutagenesis studies. These latter five mutants were tested in a murine renal abscess model, and one mutant harboring an insertion in nagD exhibited attenuated virulence. Interestingly, Tn917 was shown to have a very strong bias for insertions near the terminus of DNA replication.

  10. Development and characterization of mouse monoclonal antibodies specific for chicken interleukin 18

    Science.gov (United States)

    Four mouse monoclonal antibodies (mAbs) which are specific for chicken interleukin 18 (chIL18) were produced and characterized by enzyme-linked immunosorbent assay (ELISA), Western blotting, quantitative real-time PCR and neutralization assays. Monoclonal antibodies specific for chIL18 identified a ...

  11. Production and characterisation of monoclonal antibodies against native and disassembled human catalase

    NARCIS (Netherlands)

    Wiemer, E. A.; Ofman, R.; Middelkoop, E.; de Boer, M.; Wanders, R. J.; Tager, J. M.

    1992-01-01

    Catalase isolated from human erythrocytes was used to immunise mice, in order to generate hybridomas producing specific monoclonal antibodies to the enzyme. Hybridomas secreting anti-(catalase) antibodies were identified by a modified enzyme-linked immunosorbent assay (ELISA) using either

  12. A synthetic glycan microarray enables epitope mapping of plant cell wall glycan-directed antibodies

    DEFF Research Database (Denmark)

    Ruprecht, Colin; Bartetzko, Max P; Senf, Deborah

    2017-01-01

    In the last three decades, more than 200 monoclonal antibodies have been raised against most classes of plant cell wall polysaccharides by different laboratories world-wide. These antibodies are widely used to identify differences in plant cell wall components in mutants, organ and tissue types, ...

  13. Antibodies: an alternative for antibiotics?

    Science.gov (United States)

    Berghman, L R; Abi-Ghanem, D; Waghela, S D; Ricke, S C

    2005-04-01

    In 1967, the success of vaccination programs, combined with the seemingly unstoppable triumph of antibiotics, prompted the US Surgeon General to declare that "it was time to close the books on infectious diseases." We now know that the prediction was overly optimistic and that the fight against infectious diseases is here to stay. During the last 20 yr, infectious diseases have indeed made a staggering comeback for a variety of reasons, including resistance against existing antibiotics. As a consequence, several alternatives to antibiotics are currently being considered or reconsidered. Passive immunization (i.e., the administration of more or less pathogen-specific antibodies to the patient) prior to or after exposure to the disease-causing agent is one of those alternative strategies that was almost entirely abandoned with the introduction of chemical antibiotics but that is now gaining interest again. This review will discuss the early successes and limitations of passive immunization, formerly referred to as "serum therapy," the current use of antibody administration for prophylaxis or treatment of infectious diseases in agriculture, and, finally, recent developments in the field of antibody engineering and "molecular farming" of antibodies in various expression systems. Especially the potential of producing therapeutic antibodies in crops that are routine dietary components of farm animals, such as corn and soy beans, seems to hold promise for future application in the fight against infectious diseases.

  14. Neonatal alloimmune thrombocytopenia caused by human leucocyte antigen-B27 antibody.

    Science.gov (United States)

    Thude, H; Schorner, U; Helfricht, C; Loth, M; Maak, B; Barz, D

    2006-04-01

    Neonatal alloimmune thrombocytopenia (NAIT) occurs when maternal alloantibodies to antigens presented on foetal platelets cause their immune destruction. Whether human leucocyte antigen (HLA) antibodies can cause NAIT is controversial. Here, a patient was described who suffered from a NAIT caused by an HLA-B27 antibody. Sera from the mother and the newborn were tested for human platelet antigen antibodies and HLA antibodies by monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay, solid phase-linked immunosorbent assay (ELISA), lymphocytotoxicity assay (LCT) and flow cytometric analysis. No antibodies against cluster designation (CD)109 and platelet glycoproteins of the father were found in patient's and mother's serum. However, HLA ELISA was used to identify HLA antibody in both sera. The antibody was specified as HLA-B27 antibody. Typing results showed that the father descended HLA-B27 antigen on patient and his brother. The mother was HLA-B27 negative. It is most conceivable that the previous pregnancy of the mother induced the production of anti-HLA-B27 antibody, which crossed the placenta and subsequently caused an NAIT in the case presented.

  15. Productive common light chain libraries yield diverse panels of high affinity bispecific antibodies

    Science.gov (United States)

    Van Blarcom, Thomas; Melton, Zea; Cheung, Wai Ling; Wagstrom, Chris; McDonough, Dan; Valle Oseguera, Cendy; Ding, Sheng; Rossi, Andrea; Potluri, Shobha; Sundar, Purnima; Sirota, Marina; Yan, Yu; Jones, Jeffrey; Roe-Zurz, Zygy; Srivatsa Srinivasan, Surabhi; Zhai, Wenwu; Pons, Jaume; Rajpal, Arvind; Chaparro-Riggers, Javier

    2018-01-01

    ABSTRACT The commercial success of bispecific antibodies generally has been hindered by the complexities associated with generating appropriate molecules for both research scale and large scale manufacturing purposes. Bispecific IgG (BsIgG) based on two antibodies that use an identical common light chain can be combined with a minimal set of Fc mutations to drive heavy chain heterodimerization in order to address these challenges. However, the facile generation of common light chain antibodies with properties similar to traditional monoclonal antibodies has not been demonstrated and they have only been used sparingly. Here, we describe the design of a synthetic human antibody library based on common light chains to generate antibodies with biochemical and biophysical properties that are indistinguishable to traditional therapeutic monoclonal antibodies. We used this library to generate diverse panels of well-behaved, high affinity antibodies toward a variety of epitopes across multiple antigens, including mouse 4-1BB, a therapeutically important T cell costimulatory receptor. Over 200 BsIgG toward 4-1BB were generated using an automated purification method we developed that enables milligram-scale production of BsIgG. This approach allowed us to identify antibodies with a wide range of agonistic activity that are being used to further investigate the therapeutic potential of antibodies targeting one or more epitopes of 4-1BB. PMID:29227213

  16. Anti-carbamylated Protein Antibody Levels Correlate with Anti-Sa (Citrullinated Vimentin) Antibody Levels in Rheumatoid Arthritis.

    Science.gov (United States)

    Challener, Gregory J; Jones, Jonathan D; Pelzek, Adam J; Hamilton, B JoNell; Boire, Gilles; de Brum-Fernandes, Artur José; Masetto, Ariel; Carrier, Nathalie; Ménard, Henri A; Silverman, Gregg J; Rigby, William F C

    2016-02-01

    The presence of anticitrullinated protein antibodies (ACPA) in rheumatoid arthritis (RA) indicates a breach in immune tolerance. Recent studies indicate that this breach extends to homocitrullination of lysines with the formation of anti-carbamylated protein (anti-CarP) antibodies. We analyzed the clinical and serologic relationships of anti-CarP in 2 RA cohorts. Circulating levels of immunoglobulin G anti-CarP antibodies were determined by ELISA in established (Dartmouth-Hitchcock Medical Center) and early (Sherbrooke University Hospital Center) cohorts and evaluated for anticyclic citrullinated peptide antibodies (anti-CCP), specific ACPA, and rheumatoid factor (RF) levels using the Student t test and correlation analysis. We identified elevated anti-CarP antibodies titers in 47.0% of seropositive patients (Dartmouth, n = 164), with relationships to anti-CCP (p < 0.0001) and IgM-RF (p = 0.001). Similarly, 38.2% of seropositive patients from the Sherbrooke cohort (n = 171) had elevated anti-CarP antibodies; titers correlated to anti-CCP (p = 0.01) but not IgM-RF (p = 0.09). A strong correlation with anti-Sa was observed: 47.9% anti-Sa+ patients were anti-CarP antibodies+ versus only 25.4% anti-Sa- in the Sherbrooke cohort (p = 0.0002), and 62.6% anti-Sa+ patients versus 26.9% anti-Sa- were anti-CarP antibodies+ in Dartmouth (p < 0.0001). We found a more variable response for reactivity to citrullinated fibrinogen or to citrullinated peptides from fibrinogen and α enolase. In 2 North American RA cohorts, we observed a high prevalence of anti-CarP antibody positivity. We also describe a surprising and unexpected association of anti-CarP with anti-Sa antibodies that could not be explained by cross-reactivity. Further, considerable heterogeneity exists between anti-CarP reactivity and other citrullinated peptide reactivity, raising the question of how the pathogenesis of antibody responses for carbamylated proteins and citrullinated proteins may be linked in vivo.

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

    Directory of Open Access Journals (Sweden)

    Ickwon Choi

    2015-04-01

    Full Text Available 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.

  18. Automated Antibody De Novo Sequencing and Its Utility in Biopharmaceutical Discovery

    Science.gov (United States)

    Sen, K. Ilker; Tang, Wilfred H.; Nayak, Shruti; Kil, Yong J.; Bern, Marshall; Ozoglu, Berk; Ueberheide, Beatrix; Davis, Darryl; Becker, Christopher

    2017-05-01

    Applications of antibody de novo sequencing in the biopharmaceutical industry range from the discovery of new antibody drug candidates to identifying reagents for research and determining the primary structure of innovator products for biosimilar development. When murine, phage display, or patient-derived monoclonal antibodies against a target of interest are available, but the cDNA or the original cell line is not, de novo protein sequencing is required to humanize and recombinantly express these antibodies, followed by in vitro and in vivo testing for functional validation. Availability of fully automated software tools for monoclonal antibody de novo sequencing enables efficient and routine analysis. Here, we present a novel method to automatically de novo sequence antibodies using mass spectrometry and the Supernovo software. The robustness of the algorithm is demonstrated through a series of stress tests.

  19. Bio-preservative effect of the essential oil of the endemic Mentha piperita used alone and in combination with BacTN635 in stored minced beef meat.

    Science.gov (United States)

    Smaoui, Slim; Hsouna, Anis Ben; Lahmar, Aida; Ennouri, Karim; Mtibaa-Chakchouk, Ahlem; Sellem, Imen; Najah, Soumaya; Bouaziz, Mohamed; Mellouli, Lotfi

    2016-07-01

    The major compounds in Mentha piperita essential oil (EOMP) were menthol (33.59%) and iso-menthone (33%). The biopreservative effect of EOMP used alone at 0.25 or 0.5% and in combination with the semi-purified bacteriocin BacTN635 at 500 or 1000AU/g, on minced beef meat was evaluated by microbiological, physicochemical and sensory analyses during storage at 4°C for 21days. EOMP used alone limited the microbial deterioration of minced meat (Pmeat beef by approximately 7days. On the basis of these results, physicochemical and sensorial parameters could be used for constructing regression models to predict overall acceptability. Overall, the strongest preservative effect was achieved by using the combination of EOMP at 0.5% with BacTN535 at 1000AU/g. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Neutron scattering measurements of spatially anisotropic magnetic exchange interactions in semiconducting K0.85 Fe1.54Se2 (TN = 280 K.

    Science.gov (United States)

    Zhao, Jun; Shen, Yao; Birgeneau, R J; Gao, Miao; Lu, Zhong-Yi; Lee, D-H; Lu, X Z; Xiang, H J; Abernathy, D L; Zhao, Y

    2014-05-02

    We use neutron scattering to study the spin excitations associated with the stripe antiferromagnetic order in semiconducting K(0.85)Fe(1.54)Se(2) (T(N) = 280 K). We show that the spin-wave spectra can be accurately described by an effective Heisenberg Hamiltonian with highly anisotropic inplane couplings at T = 5 K. At high temperature (T = 300 K) above T(N), short-range magnetic correlation with anisotropic correlation lengths are observed. Our results suggest that, despite the dramatic difference in the Fermi surface topology, the inplane anisotropic magnetic couplings are a fundamental property of the iron-based compounds; this implies that their antiferromagnetism may originate from local strong correlation effects rather than weak coupling Fermi surface nesting.

  1. Relevance of hot spots in the evolution and transmission of Tn1546 in glycopeptide-resistant Enterococcus faecium (GREF) from broiler origin

    DEFF Research Database (Denmark)

    Migura, Lourdes Garcia; Hasman, Henrik; Svendsen, Christina Aaby

    2008-01-01

    found in isolates from the UK (n = 150). The first insertion point revealed that 25 isolates harboured Tn1546 positioned in a sequence with 96% homology to a streptomycin adenyltransferase gene (AY604739) from a Staphylococcus intermedius plasmid. At this insertion point, a direct repeat (GTCCT......) was duplicated as previously described, indicating transposition at the target site. Furthermore, this 'hot spot' was also detected in isolates from Norway (2/8) and Denmark (17/20). The second insertion point detected in 45 isolates from the UK revealed integration into an Inc18-like plasmid, most likely...... by a process of target site recombination. Conclusions: The presence of a common insertion point for isolates from different geographical areas could suggest the insertion of Tn1546 by transposition in a plasmid-specific site, followed by genetic rearrangement both inside the transposon and in the flanking...

  2. CREATINE PHOSPHOKINASE ISOENZYME-MB MASS (CK-MB MASS AND TROPONIN I (cTnI IN DOGS (Canis familiaris

    Directory of Open Access Journals (Sweden)

    Millena Vidal Freitas

    2015-07-01

    Full Text Available Cardiac markers, especially CK-MB mass and cTnI, have an essential role in both human and veterinary clinical and surgical cardiology, allowing a more precise and accurate diagnosis in myocardial lesions. The goal of this work was to measure these cardiac markers in veterinary medicine, improve their use and provide information about these laboratory methods that allow non-invasive health monitoring of the myocardial cell. Parameters quantification was obtained from sera of healthy dogs examined during routine procedures at the Small Animal Veterinary Hospital of Universidade Estadual do Norte Fluminense Darcy Ribeiro. The human chemiluminescence essay turbo kit (IMMULITE Turbo, Siemens ® test proved to be effective in canine species for both CK-MB mass and cTnI. In addition, the values found for CK-MB will significantly contribute to clinical routine or experimental work with dogs.

  3. Recent Approaches to Modeling Transport of Mercury in Surface Water and Groundwater - Case Study in Upper East Fork Poplar Creek, Oak Ridge, TN - 13349

    International Nuclear Information System (INIS)

    Bostick, Kent; Daniel, Anamary; Tachiev, Georgio; Malek-Mohammadi, Siamak

    2013-01-01

    In this case study, groundwater/surface water modeling was used to determine efficacy of stabilization in place with hydrologic isolation for remediation of mercury contaminated areas in the Upper East Fork Poplar Creek (UEFPC) Watershed in Oak Ridge, TN. The modeling simulates the potential for mercury in soil to contaminate groundwater above industrial use risk standards and to contribute to surface water contamination. The modeling approach is unique in that it couples watershed hydrology with the total mercury transport and provides a tool for analysis of changes in mercury load related to daily precipitation, evaporation, and runoff from storms. The model also allows for simulation of colloidal transport of total mercury in surface water. Previous models for the watershed only simulated average yearly conditions and dissolved concentrations that are not sufficient for predicting mercury flux under variable flow conditions that control colloidal transport of mercury in the watershed. The transport of mercury from groundwater to surface water from mercury sources identified from information in the Oak Ridge Environmental Information System was simulated using a watershed scale model calibrated to match observed daily creek flow, total suspended solids and mercury fluxes. Mercury sources at the former Building 81-10 area, where mercury was previously retorted, were modeled using a telescopic refined mesh with boundary conditions extracted from the watershed model. Modeling on a watershed scale indicated that only source excavation for soils/sediment in the vicinity of UEFPC had any effect on mercury flux in surface water. The simulations showed that colloidal transport contributed 85 percent of the total mercury flux leaving the UEFPC watershed under high flow conditions. Simulation of dissolved mercury transport from liquid elemental mercury and adsorbed sources in soil at former Building 81-10 indicated that dissolved concentrations are orders of magnitude

  4. Oligopeptide-based enzyme immunoassay for ovine lentivirus antibody detection.

    OpenAIRE

    Kwang, J; Torres, J V

    1994-01-01

    Ovine progressive pneumonia virus (OPPV) is a lentivirus which causes a progressive disease in sheep. Immunodominant epitopes have been identified in the envelope gp40 glycoprotein. Synthetic peptides representing these regions are able to detect the presence of OPPV antibodies in 96% of infected sheep.

  5. Discrimination between Native and Tn6010-Associated oqxAB in Klebsiella spp., Raoultella spp., and other Enterobacteriaceae by Using a Two-Step Strategy

    DEFF Research Database (Denmark)

    Guillard, Thomas; Lebreil, Anne-Laure; Hansen, Lars Hestbjerg

    2015-01-01

    ABSTRACT We developed a two-step PCR-based strategy to detect genes encoding OqxAB, allowing a specific assignment of Tn6010-associated oqxAB in Enterobacteriaceae. Chromosomal location in this setup was confirmed by hybridization with I-CeuI-restricted genomes. This approach led us to find that ...... that Klebsiella sp. and Raoultella sp. reference strains chromosomally carried oqxAB....

  6. Zbiornik w wyrobisku końcowym Odkrywki "Pątnów" Kopalni Węgla Brunatnego "Konin" i jego bilans wodny za okres 2003-2004

    OpenAIRE

    Wachowiak, Grzegorz; Wachowiak, Agata

    2005-01-01

    The idea of water recultivation and management of the closing open pits in the Lignite Opencast Mine "Konin" is of a growing importance. Hence, the necessity to explore water relations conditioned by SLich undertakings. Recultivation scheme of the closing open pit "Pątnów" of Lignite Opencast Mine "Konin" includes the construction of a 346 ha reservoir with the capacity of over 38 mln m. So far, it is the biggest reservoir built as a result of an open pit reculti...

  7. Current perspectives on antibody-mediated rejection after lung transplantation

    Directory of Open Access Journals (Sweden)

    Witt CA

    2014-10-01

    Full Text Available Chad A Witt, Ramsey R Hachem Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, Washington University School of Medicine, Saint Louis, MO, USA Abstract: The role of donor-specific antibodies (DSA to human leukocyte antigens and the burden of antibody-mediated rejection (AMR in lung transplantation remain enigmatic. Over the past several years, evidence has been emerging that humoral immunity plays an important role in the development of both acute and chronic lung allograft dysfunction (CLAD. Multiple case reports and case series have identified lung allograft recipients with clinical findings consistent with acute AMR. However, there is currently no widely accepted definition for AMR in lung transplantation, and this has been a significant barrier to furthering our understanding of this form of rejection. Nonetheless, the development of DSA after transplantation has consistently been identified as an independent risk factor for persistent and high-grade acute cellular rejection and CLAD. This has raised the possibility that chronic AMR may be a distinct phenotype of CLAD although evidence supporting this paradigm is still lacking. Additionally, antibodies to lung-restricted self-antigens (collagen V and K-α 1 tubulin have been associated with primary graft dysfunction early and the development of CLAD late after transplantation, and emerging evidence underscores significant interactions between autoimmunity and alloimmunity after transplantation. There is currently an active International Society for Heart and Lung Transplantation working group that is developing an operational definition for AMR in lung transplantation. This will be critical to improve our understanding of this form of rejection and conduct clinical trials to identify optimal treatment strategies. This review will summarize the literature on DSA and AMR in lung transplantation and discuss the impact of antibodies to self-antigens on lung

  8. A simple and novel method for RNA-seq library preparation of single cell cDNA analysis by hyperactive Tn5 transposase.

    Science.gov (United States)

    Brouilette, Scott; Kuersten, Scott; Mein, Charles; Bozek, Monika; Terry, Anna; Dias, Kerith-Rae; Bhaw-Rosun, Leena; Shintani, Yasunori; Coppen, Steven; Ikebe, Chiho; Sawhney, Vinit; Campbell, Niall; Kaneko, Masahiro; Tano, Nobuko; Ishida, Hidekazu; Suzuki, Ken; Yashiro, Kenta

    2012-10-01

    Deep sequencing of single cell-derived cDNAs offers novel insights into oncogenesis and embryogenesis. However, traditional library preparation for RNA-seq analysis requires multiple steps with consequent sample loss and stochastic variation at each step significantly affecting output. Thus, a simpler and better protocol is desirable. The recently developed hyperactive Tn5-mediated library preparation, which brings high quality libraries, is likely one of the solutions. Here, we tested the applicability of hyperactive Tn5-mediated library preparation to deep sequencing of single cell cDNA, optimized the protocol, and compared it with the conventional method based on sonication. This new technique does not require any expensive or special equipment, which secures wider availability. A library was constructed from only 100 ng of cDNA, which enables the saving of precious specimens. Only a few steps of robust enzymatic reaction resulted in saved time, enabling more specimens to be prepared at once, and with a more reproducible size distribution among the different specimens. The obtained RNA-seq results were comparable to the conventional method. Thus, this Tn5-mediated preparation is applicable for anyone who aims to carry out deep sequencing for single cell cDNAs. Copyright © 2012 Wiley Periodicals, Inc.

  9. Effect of PCI on inflammatory factors, cTnI, MMP-9 and NT-pro BNP in patients with unstable angina pectoris

    Directory of Open Access Journals (Sweden)

    Ke-Tong Liu

    2016-05-01

    Full Text Available Objective: To investigate the effect of PCI on inflammatory factors, cTnI, MMP-9and NTpro BNP in patients with unstable angina pectoris. Methods: A total of 80 unstable angina pectoris patients were divided into observation group (40 cases and control group (40 cases. The observation group was given the therapy of PCI, and the control group was given coronary angiography. To observe the of inflammatory factors, cTnI, MMP-9 and NT-pro BNP were tested and compared before and after operation. Results: At 24 h after operation, CRP and IL-18 levels were increased significantly after treatment inoperation groups, there was no difference on inflammatory factors in control group, and had significant difference on inflammatory factors in two groups; At 24 h after operation, cTnI, MMP-9 and NT-pro BNP levels were increased significantly after treatment inoperation groups, there was no difference on inflammatory factors in control group, and had significant difference on inflammatory factors in two groups. Conclusion: PCI therapy can induce inflammation and myocardial injury in patients with unstable angina pectoris.

  10. Hfq restructures RNA-IN and RNA-OUT and facilitates antisense pairing in the Tn10/IS10 system

    Science.gov (United States)

    Ross, Joseph A.; Ellis, Michael J.; Hossain, Shahan; Haniford, David B.

    2013-01-01

    Hfq functions in post-transcriptional gene regulation in a wide range of bacteria, usually by promoting base-pairing of mRNAs and trans-encoded sRNAs that share partial sequence complementarity. It is less clear if Hfq is required for pairing of cis-encoded RNAs (i.e., antisense RNAs) with their target mRNAs. In the current work, we have characterized the interactions between Escherichia coli Hfq and the components of the Tn10/IS10 antisense system, RNA-IN and RNA-OUT. We show that Hfq interacts with RNA-OUT through its proximal RNA-binding surface, as is typical for Hfq and trans-encoded sRNAs. In contrast, RNA-IN binds both proximal and distal RNA-binding surfaces in Hfq with a higher affinity for the latter, as is typical for mRNA interactions in canonical sRNA-mRNA pairs. Importantly, an amino acid substitution in Hfq that interferes with RNA binding to the proximal site negatively impacts RNA-IN:OUT pairing in vitro and suppresses the ability of Hfq to negatively regulate IS10 transposition in vivo. We also show that Hfq binding to RNA-IN and RNA-OUT alters secondary structure elements in both of these RNAs and speculate that this could be important in how Hfq facilitates RNA-IN:OUT pairing. Based on the results presented here, we suggest that Hfq could be involved in regulating RNA pairing in other antisense systems, including systems encoded by other transposable elements. PMID:23510801

  11. Time dependence of volcano inflation: mass influx or viscoelastic relaxation? Insights from Grímsvötn volcano, Iceland

    Science.gov (United States)

    Segall, P.

    2017-12-01

    Distinguishing magma chamber pressurization from relaxation of a viscoelastic aureole surrounding the chamber based on geodetic measurements has remained challenging. Elastic models with mass inflow proportional to the pressure difference between the chamber and a deep reservoir predict exponentially decaying flux. For a spherical chamber surrounded by a Maxwell viscoelastic shell with pressure dependent recharge, the surface deformation is the sum of two exponentials (Segall, 2016). GPS displacements following eruptions of Grímsvötn, Iceland in 2004 and 2011 exhibit rapid post-eruptive inflation (time scale of 0.1 yr), followed by inflation with a much longer time constant. Markov Chain Monte Carlo inversion with the viscoelastic model shows the GPS time series can be fit with viscosity of 2e16 Pa-s, and a relatively incompressible magma, B = beta_c/ (beta_m + beta_c) > 0.6, where beta_m and beta_c are chamber and magma compressibility. The latter appears to conflict with the ratio of erupted volume to geodetically inferred source volume change, rv 10, obtained for the best fitting spherical (Mogi ) source (Hreinsdóttir, 2014). Since rv = 1/B, this implies a relatively compressible melt, B 0.1. Reexamination of the co-eruptive GPS and tilt data with the more general ellipsoidal model of Cervelli (2013), reveals that the best fitting sources are oblate (b/a 3), deeper, and with larger volume changes, rv 3, relative to spherical models. Oblate magma chambers are consistent with seismic tomography. FEM calculations including free surface effects lead to even larger co-eruptive volume changes, smaller rv and hence larger B. I conclude that the data are consistent with rapid post-eruptive inflation driven by viscoelastic relaxation with a relatively incompressible magma, although other interpretations will be discussed.

  12. Hu and Yo antibodies have heterogeneous avidity.

    Science.gov (United States)

    Totland, Cecilie; Aarseth, Jan; Vedeler, Christian

    2007-04-01

    Onconeural antibodies such as anti-Hu and anti-Yo may be important in the pathogenesis of paraneoplastic neurological syndromes. The avidity of these antibodies is not known. In this study, we compared the avidity of Hu and Yo antibodies both at single time points and over a time range of 2 months to 6 years. The avidity of Yo and Hu antibodies differed among the patients, but anti-Yo generally had higher avidity than anti-Hu. Whether Yo antibodies are more pathogenic than Hu antibodies are presently unknown.

  13. Detection and differentiation of cytomegalovirus antibodies by radioimmunoassay

    International Nuclear Information System (INIS)

    Jankowski, M.A.; Gut, W.; Nawrocka, E.; Kantoch, M.

    1980-01-01

    A sensitive radioimmunological method was adapted for the assay of anticytomegalovirus IgG and IgM antibodies. Binding of immunoglobulins G with cytomegalovirus (CMV) antigens was greatly reduced when an antigen prepared by sonication of the infected cells in glycine buffer was used in place of the antigen obtained by freezing and thawing of the cells. The application of labelled serum against the Fc fragment of IgG made it possible to identify selectively the antibodies bound with the virus antigen by antigenic determinants. (Auth.)

  14. HYDROLOGY, SUMNER COUNTY, TN

    Data.gov (United States)

    Federal Emergency Management Agency, Department of Homeland Security — Recent developments in digital terrain and geospatial database management technology make it possible to protect this investment for existing and future projects to...

  15. Conformational Occlusion of Blockade Antibody Epitopes, a Novel Mechanism of GII.4 Human Norovirus Immune Evasion.

    Science.gov (United States)

    Lindesmith, Lisa C; Mallory, Michael L; Debbink, Kari; Donaldson, Eric F; Brewer-Jensen, Paul D; Swann, Excel W; Sheahan, Timothy P; Graham, Rachel L; Beltramello, Martina; Corti, Davide; Lanzavecchia, Antonio; Baric, Ralph S

    2018-01-01

    Extensive antigenic diversity within the GII.4 genotype of human norovirus is a major driver of pandemic emergence and a significant obstacle to development of cross-protective immunity after natural infection and vaccination. However, human and mouse monoclonal antibody studies indicate that, although rare, antibodies to conserved GII.4 blockade epitopes are generated. The mechanisms by which these epitopes evade immune surveillance are uncertain. Here, we developed a new approach for identifying conserved GII.4 norovirus epitopes. Utilizing a unique set of virus-like particles (VLPs) representing the in vivo -evolved sequence diversity within an immunocompromised person, we identify key residues within epitope F, a conserved GII.4 blockade antibody epitope. The residues critical for antibody binding are proximal to evolving blockade epitope E. Like epitope F, antibody blockade of epitope E was temperature sensitive, indicating that particle conformation regulates antibody access not only to the conserved GII.4 blockade epitope F but also to the evolving epitope E. These data highlight novel GII.4 mechanisms to protect blockade antibody epitopes, map essential residues of a GII.4 conserved epitope, and expand our understanding of how viral particle dynamics may drive antigenicity and antibody-mediated protection by effectively shielding blockade epitopes. Our data support the notion that GII.4 particle breathing may well represent a major mechanism of humoral immune evasion supporting cyclic pandemic virus persistence and spread in human populations. IMPORTANCE In this study, we use norovirus virus-like particles to identify key residues of a conserved GII.4 blockade antibody epitope. Further, we identify an additional GII.4 blockade antibody epitope to be occluded, with antibody access governed by temperature and particle dynamics. These findings provide additional support for particle conformation-based presentation of binding residues mediated by a particle

  16. [Immunohematologic study and transfusion approach to patients with public antibodies].

    Science.gov (United States)

    Solves, P; de la Rubia, J; Arriaga, F; Cervera, J; Arnao, M; Carpio, N; Marty, M L

    1997-02-01

    To analyze the different immunohematologic studies required to identify anti-red cell antibodies directed against high incidence antigens and comment the best tranfusion management. Five patients with suspected anti-red cell alloantibodies directed against high frequency antigens are reported. After a positive antibody screening test (AST), an agglutination test with a commercial panel of 24 red cells was performed. Red cells were treated with proteolytic enzymes and AET to try to identify the circulating antibody. However, it was necessary to send the samples to reference laboratories for definitive identification. In order to evaluate the haemolytic potential of the antibody serum samples were treated with DTT and immunoglobulin subtype was studied with the capillary agglutination test. Finally, we analyze the half life of Cr51 labelled red cells. To obtain compatible blood for transfusion, autologous transfusion and cross-match with blood from direct relatives were performed. AST was positive in every case. A decrease in the agglutination test was observed after ficin treatment in two patients, and an increase in the remaining. The treatment of red cells with ZZAP and AET resulted in a decrease of agglutination in three cases and an increase in the remaining two. Specificity of the antibodies was as follows: anti-Cellano (two cases), anti-Ku (one case) and anti-Yta (two cases). Anti-Kell antibodies were IgG1 and anti-Cartwright antibodies were IgG4. One patient was transfused with autologous blood alone, another patient received compatible blood from direct relatives. A third patient was transfused both with autologous and allogeneic compatible blood. The fourth patient did not need red cell transfusion and, finally the last patient had to be transfused with incompatible blood but no postransfusion haemolysis was observed. In patients with anti-red cell antibodies against high-frequency antigens, red blood cells treatment with proteolytic enzymes (ZZAP, ficin

  17. Glycine receptor antibodies are detected in progressive encephalomyelitis with rigidity and myoclonus (PERM) but not in saccadic oscillations.

    Science.gov (United States)

    Iizuka, Takahiro; Leite, Maria I; Lang, Bethan; Waters, Patrick; Urano, Yoshiaki; Miyakawa, Saori; Hamada, Junichi; Sakai, Fumihiko; Mochizuki, Hideki; Vincent, Angela

    2012-08-01

    Glycine receptor (GlyR) antibodies were recently identified in a few patients with progressive encephalomyelitis with rigidity and myoclonus (PERM); none of these patients had antibodies against glutamic acid decarboxylase (GAD). An inhibitory glycinergic transmission defect has also been implicated in the mechanism underlying saccadic oscillations, including ocular flutter or opsoclonus; GlyR antibodies have not been reported in these patients. The purpose was to determine whether GlyR antibodies are found in patients with PERM, ocular flutter syndrome (OFS), and opsoclonus-myoclonus syndrome (OMS). GlyR antibodies were first measured in archived sera and CSF from five patients, including one patient with GAD antibody-positive PERM, two patients with OFS, and two patients with OMS. GlyR antibodies were also measured in archived sera from nine other adult patients with OMS. GlyR antibodies and GAD antibodies were both found at high titers in the serum and CSF of the patient with PERM, and their levels paralleled disease activity over time. GlyR antibodies were not found at significant levels in 13 patients with saccadic oscillations. GlyR and GAD antibodies can co-exist in PERM and follow the clinical course. Although saccadic oscillations are a feature of this condition, GlyR antibodies are not commonly found in patients with isolated saccadic oscillations.

  18. Prediction of the Hydrogen Peroxide-Induced Methionine Oxidation Propensity in Monoclonal Antibodies.

    Science.gov (United States)

    Agrawal, Neeraj J; Dykstra, Andrew; Yang, Jane; Yue, Hai; Nguyen, Xichdao; Kolvenbach, Carl; Angell, Nicolas

    2018-01-08

    Methionine oxidation in therapeutic antibodies can impact the product's stability, clinical efficacy, and safety and hence it is desirable to address the methionine oxidation liability during antibody discovery and development phase. Although the current experimental approaches can identify the oxidation-labile methionine residues, their application is limited mostly to the development phase. We demonstrate an in silico method that can be used to predict oxidation-labile residues based solely on the antibody sequence and structure information. Since antibody sequence information is available in the discovery phase, the in silico method can be applied very early on to identify the oxidation-labile methionine residues and subsequently address the oxidation liability. We believe that the in silico method for methionine oxidation liability assessment can aid in antibody discovery and development phase to address the liability in a more rational way. Copyright © 2018 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  19. 9 CFR 113.452 - Erysipelothrix Rhusiopathiae Antibody.

    Science.gov (United States)

    2010-01-01

    ..., DEPARTMENT OF AGRICULTURE VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS STANDARD... Antibody is a specific antibody product containing antibodies directed against one or more somatic antigens...

  20. HER2 monoclonal antibodies that do not interfere with receptor heterodimerization-mediated signaling induce effective internalization and represent valuable components for rational antibody-drug conjugate design.

    Science.gov (United States)

    de Goeij, Bart E C G; Peipp, Matthias; de Haij, Simone; van den Brink, Edward N; Kellner, Christian; Riedl, Thilo; de Jong, Rob; Vink, Tom; Strumane, Kristin; Bleeker, Wim K; Parren, Paul W H I

    2014-01-01

    The human epidermal growth factor receptor (HER)2 provides an excellent target for selective delivery of cytotoxic drugs to tumor cells by antibody-drug conjugates (ADC) as has been clinically validated by ado-trastuzumab emtansine (Kadcyla(TM)). While selecting a suitable antibody for an ADC approach often takes specificity and efficient antibody-target complex internalization into account, the characteristics of the optimal antibody candidate remain poorly understood. We studied a large panel of human HER2 antibodies to identify the characteristics that make them most suitable for an ADC approach. As a model toxin, amenable to in vitro high-throughput screening, we employed Pseudomonas exotoxin A (ETA') fused to an anti-kappa light chain domain antibody. Cytotoxicity induced by HER2 antibodies, which were thus non-covalently linked to ETA', was assessed for high and low HER2 expressing tumor cell lines and correlated with internalization and downmodulation of HER2 antibody-target complexes. Our results demonstrate that HER2 antibodies that do not inhibit heterodimerization of HER2 with related ErbB receptors internalize more efficiently and show greater ETA'-mediated cytotoxicity than antibodies that do inhibit such heterodimerization. Moreover, stimulation with ErbB ligand significantly enhanced ADC-mediated tumor kill by antibodies that do not inhibit HER2 heterodimerization. This suggests that the formation of HER2/ErbB-heterodimers enhances ADC internalization and subsequent killing of tumor cells. Our study indicates that selecting HER2 ADCs that allow piggybacking of HER2 onto other ErbB receptors provides an attractive strategy for increasing ADC delivery and tumor cell killing capacity to both high and low HER2 expressing tumor cells.

  1. Anti-Ro52 antibody level is an important marker of fetal congenital heart block risk in anti-Ro/SSA antibody positive pregnancy.

    Science.gov (United States)

    Miyasato-Isoda, Mai; Waguri, Masako; Yamada, Yuko; Miyano, Akira; Wada, Yoshinao

    2017-09-21

    The aims of this study are to determine the incidence of congenital heart block (CHB) in the Japanese population and identify maternal factors predicting fetal CHB in anti-Ro/SSA antibody positive pregnancy. A retrospective study was performed using 52,147 clinical records of pregnancies followed in a single center. For 183 anti-Ro/SSA antibody-positive women, anti-Ro52 and Ro60 antibodies were measured, and the odds of CHB in relation to maternal clinical features were calculated by multivariate analysis. The receiver-operating characteristic (ROC) curves for predicting CHB were constructed for the titers of anti-Ro/SSA, anti-Ro52 and anti-Ro60 antibodies. Fetal CHB occurred in two pregnancies among those without known risks such as positive anti-Ro/SSA antibody or previous CHB-affected pregnancy, suggesting an incidence similar to that in Caucasian populations. As for the anti-Ro/SSA antibody positive pregnancies, the titers of anti-Ro/SSA, anti-Ro52 and anti-Ro60 antibodies were independent risk factors for fetal CHB and the use of corticosteroids before 18 gestational weeks was an independent protective factor. The area under the ROC was 0.84, 0.73 and 0.74 for anti-Ro52, anti-Ro60 and anti-Ro/SSA antibodies, respectively. CHB occurred in two among approximately 50,000 pregnancies without known risks such as positive anti-Ro/SSA antibody or previous delivery of CHB-affected babies. Measurement of anti-Ro52 antibody levels may be helpful in extracting a risk group of delivering CHB infants in the anti-Ro/SSA antibody positive pregnancy.

  2. Marrow Derived Antibody Library for the Treatment of Neuroblastoma

    Science.gov (United States)

    2015-12-01

    identified with strong binding to tumor cells. In Figure 1, the positive binding of phage clones to MGT-003 cells were visualized using anti- M13 phage ...neuroblastoma patients using phage display and B cell hybridoma technologies. The scope of this project is to use NB patient-derived materials to...create NB cell lines, xenograft models, NB specific phage display libraries and to identify and amplify functional anti-NB specific antibodies for future

  3. Monoclonal antibodies to Treponema Pallidum.

    NARCIS (Netherlands)

    H.J.M. van de Donk; J.D.A. van Embden; M.F. van Olderen; A.D.M.E. Osterhaus (Albert); J.C. de Jong (Jan)

    1984-01-01

    textabstractThree successive fusions of mouse myeloma cells and spleen lymphocytes of a mouse immunized with Treponema Pallidum resulted in one hybridoma producing anti T. pallidum antibodies for each fusion. The mice were immunized with live pallidum cells respectively 1, 3 and 5 months before

  4. [Detection of antibodies to mycoplasmas using an immunoenzyme method].

    Science.gov (United States)

    Hájková, M; Jurmanová, K

    1986-08-01

    Detection of the antibodies to the species Mycoplasma bovis in the serum and milk of dairy cows coming from a mastitis-infected herd is a good example of utilization of the ELISA immunoenzymologic method in the mycoplasmology. Examining the samples from 75 dairy cows and applying the indirect hemagglutination test, good correlation of the results of the two tests was determined. The antibodies to the species Ureaplasma diversum were demonstrated by the ELISA method both in the bovine serum and in the milk of dairy cows infected slightly with mastitis. We chosen that strain which detected the maximum titres in the selected samples of the sera out of four antigens prepared from various strains of U. diversum. Rabbit sera hyperimmune to 26 strains of the mycoplasmas of various species were used to identify two antigens (after removing the antibodies to the components of the media). Specific reaction was obtained with the antisera to M. hyorhinis and M. arginini.

  5. Antibody induction versus corticosteroid induction for liver transplant recipients

    DEFF Research Database (Denmark)

    Penninga, Luit; Wettergren, André; Wilson, Colin H

    2014-01-01

    . OBJECTIVES: To assess the benefits and harms of T-cell specific antibody induction versus corticosteroid induction for prevention of acute rejection in liver transplant recipients. SEARCH METHODS: We searched The Cochrane Hepato-Biliary Group Controlled Trials Register, the Cochrane Central Register...... to identify additional trials. SELECTION CRITERIA: We included all randomised clinical trials assessing immunosuppression with T-cell specific antibody induction versus corticosteroid induction in liver transplant recipients. Our inclusion criteria stated that participants within each included trial should...... (bias) using bias risk domains with definitions. We used trial sequential analysis to control for random errors (play of chance). MAIN RESULTS: We included 10 randomised trials with a total of 1589 liver transplant recipients, which studied the use of T-cell specific antibody induction versus...

  6. Antibodies recognizing both IgM isotypes in Atlantic salmon

    DEFF Research Database (Denmark)

    Hedfors, Ida Aagård; Bakke, Hege; Skjødt, Karsten

    2012-01-01

    these molecules. The present study aimed at identifying tools to separate IgM positive (IgM(+)) B cells from IgM negative (IgM(-)) non-B cell populations using flow cytometry. Several monoclonal antibodies (mAbs), and one polyclonal antibody (pAb) to both rainbow trout (Oncorhynchus mykiss) and Atlantic salmon...... (Salmo salar) IgM, either commercially available or locally produced were tested for their recognition of Atlantic salmon IgM(+) cells. Leukocytes were isolated from peripheral blood (PB), spleen (S) and head kidney (HK) and stained with all mAbs and the pAb, to possibly verify the approximate number...... of IgM(+) cells in the respective tissues in salmon. To our surprise, this seemingly simple task did not reveal similar staining patterns for all antibodies as expected, but rather large differences in the number of positively stained cells were discovered. In short, positively stained cells by each...

  7. Development of human antibody fragments using antibody phage display for the detection and diagnosis of Venezuelan equine encephalitis virus (VEEV

    Directory of Open Access Journals (Sweden)

    Hust Michael

    2008-09-01

    Full Text Available Abstract Background Venezuelan equine encephalitis virus (VEEV belongs to the Alphavirus group. Several species of this family are also pathogenic to humans and are recognized as potential agents of biological warfare and terrorism. The objective of this work was the generation of recombinant antibodies for the detection of VEEV after a potential bioterrorism assault or an natural outbreak of VEEV. Results In this work, human anti-VEEV single chain Fragments variable (scFv were isolated for the first time from a human naïve antibody gene library using optimized selection processes. In total eleven different scFvs were identified and their immunological specificity was assessed. The specific detection of the VEEV strains TC83, H12/93 and 230 by the selected antibody fragments was proved. Active as well as formalin inactivated virus particles were recognized by the selected antibody fragments which could be also used for Western blot analysis of VEEV proteins and immunohistochemistry of VEEV infected cells. The anti-VEEV scFv phage clones did not show any cross-reactivity with Alphavirus species of the Western equine encephalitis virus (WEEV and Eastern equine encephalitis virus (EEEV antigenic complex, nor did they react with Chikungunya virus (CHIKV, if they were used as detection reagent. Conclusion For the first time, this study describes the selection of antibodies against a human pathogenic virus from a human naïve scFv antibody gene library using complete, active virus particles as antigen. The broad and sensitive applicability of scFv-presenting phage for the immunological detection and diagnosis of Alphavirus species was demonstrated. The selected antibody fragments will improve the fast identification of VEEV in case of a biological warfare or terroristic attack or a natural outbreak.

  8. Physical, chemical and biological CTD and bottle data from R/V Thomas G. Thompson cruise TN278 in eastern tropical North Pacific Ocean from March 19 to April 20, 2012 (NODC Accession 0109846)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This report contains data from R/V Thomas G. Thompson cruise TN278 to the eastern tropical north pacific oxygen deficient zone. The objective of the cruise was to...

  9. Protective Capacity of the Human Anamnestic Antibody Response during Acute Dengue Virus Infection.

    Science.gov (United States)

    Xu, Meihui; Züst, Roland; Toh, Ying Xiu; Pfaff, Jennifer M; Kahle, Kristen M; Davidson, Edgar; Doranz, Benjamin J; Velumani, Sumathy; Tukijan, Farhana; Wang, Cheng-I; Fink, Katja

    2016-12-15

    Half of the world's population is exposed to the risk of dengue virus infection. Although a vaccine for dengue virus is now available in a few countries, its reported overall efficacy of about 60% is not ideal. Protective immune correlates following natural dengue virus infection remain undefined, which makes it difficult to predict the efficacy of new vaccines. In this study, we address the protective capacity of dengue virus-specific antibodies that are produced by plasmablasts a few days after natural secondary infection. Among a panel of 18 dengue virus-reactive human monoclonal antibodies, four groups of antibodies were identified based on their binding properties. While antibodies targeting the fusion loop of the glycoprotein of dengue virus dominated the antibody response, two smaller groups of antibodies bound to previously undescribed epitopes in domain II of the E protein. The latter, largely serotype-cross-reactive antibodies, demonstrated increased stability of binding at pH 5. These antibodies possessed weak to moderate neutralization capacity in vitro but were the most efficacious in promoting the survival of infected mice. Our data suggest that the cross-reactive anamnestic antibody response has a protective capacity despite moderate neutralization in vitro and a moderate decrease of viremia in vivo IMPORTANCE: Antibodies can protect from symptomatic dengue virus infection. However, it is not easy to assess which classes of antibodies provide protection because in vitro assays are not always predictive of in vivo protection. During a repeat infection, dengue virus-specific immune memory cells are reactivated and large amounts of antibodies are produced. By studying antibodies cloned from patients with heterologous secondary infection, we tested the protective value of the serotype-cross-reactive "recall" or "anamnestic" response. We found that results from in vitro neutralization assays did not always correlate with the ability of the antibodies to

  10. Radioimmunological determination of growth hormone antibodies

    International Nuclear Information System (INIS)

    Kracmar, P.; Hnikova, O.

    1979-01-01

    The method is based on the assumption of the presence of antibodies in the serum of the patient and the formation of the complex antibody-tracer ( 125 I-STH). For separation the principle is used of two antibodies and subsequent ultrafiltration with membrane ultrafilters. Clinical experience, reproducibility and the procedure recommended for simple monitoring and the determination of the amount of antibodies in the serum of patients are presented. (author)

  11. Antibody therapeutics - the evolving patent landscape.

    Science.gov (United States)

    Petering, Jenny; McManamny, Patrick; Honeyman, Jane

    2011-09-01

    The antibody patent landscape has evolved dramatically over the past 30 years, particularly in areas of technology relating to antibody modification to reduce immunogenicity in humans or improve antibody function. In some cases antibody techniques that were developed in the 1980s are still the subject of patent protection in the United States or Canada. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Antibodies against chromosomal beta-lactamase

    DEFF Research Database (Denmark)

    Giwercman, B; Rasmussen, J W; Ciofu, Oana

    1994-01-01

    A murine monoclonal anti-chromosomal beta-lactamase antibody was developed and an immunoblotting technique was used to study the presence of serum and sputum antibodies against Pseudomonas aeruginosa chromosomal group 1 beta-lactamase in patients with cystic fibrosis (CF). The serum antibody resp...... against the infection. On the other hand, immune complexes between the beta-lactamase and corresponding antibodies could play a role in the pathogenesis of bronchopulmonary injury in CF by mediating hyperimmune reactions....

  13. Human antibody fragments specific for the epidermal growth factor receptor selected from large non-immunised phage display libraries.

    Science.gov (United States)

    Souriau, Christelle; Rothacker, Julie; Hoogenboom, Hennie R; Nice, Edouard

    2004-09-01

    Antibodies to EGFR have been shown to display anti-tumour effects mediated in part by inhibition of cellular proliferation and angiogenesis, and by enhancement of apoptosis. Humanised antibodies are preferred for clinical use to reduce complications with HAMA and HAHA responses frequently seen with murine and chimaeric antibodies. We have used depletion and subtractive selection strategies on cells expressing the EGFR to sample two large antibody fragment phage display libraries for the presence of human antibodies which are specific for the EGFR. Four Fab fragments and six scFv fragments were identified, with affinities of up to 2.2nM as determined by BIAcore analysis using global fitting of the binding curves to obtain the individual rate constants (ka and kd). This overall approach offers a generic screening method for the identification of growth factor specific antibodies and antibody fragments from large expression libraries and has potential for the rapid development of new therapeutic and diagnostic reagents.

  14. Radioimmunoassay method for detection of gonorrhea antibodies

    International Nuclear Information System (INIS)

    1975-01-01

    A novel radioimmunoassay for the detection of gonorrhea antibodies in serum is described. A radionuclide is bound to gonorrhea antigens produced by a growth culture. In the presence of gonorrhea antibodies in the serum, an antigen-antibody conjugate is formed, the concentration of which can be measured with conventional radiometric methods. The radioimmunoassay is highly specific

  15. Antibodies Against Melanin | Wassermann | South African Medical ...

    African Journals Online (AJOL)

    This study reports on unsuccessful attempts to produce antibodies against melanoprotein in rabbits. Available evidence suggests antibodies against melanocytes in the aetiology of vitiligo, but there is no convincing evidence for antibodies against melanin per se. It is suggested that the demonstration of antibodif's against ...

  16. Selection of apoptotic cell specific human antibodies from adult bone marrow.

    Directory of Open Access Journals (Sweden)

    Caroline Grönwall

    Full Text Available Autoreactive antibodies that recognize neo-determinants on apoptotic cells in mice have been proposed to have protective, homeostatic and immunoregulatory properties, although our knowledge about the equivalent antibodies in humans has been much more limited. In the current study, human monoclonal antibodies with binding specificity for apoptotic cells were isolated from the bone marrow of healthy adults using phage display technology. These antibodies were shown to recognize phosphorylcholine (PC-associated neo-determinants. Interestingly, three of the four identified apoptotic cell-specific antibody clones were encoded by VH3 region rearrangements with germline or nearly germline configuration without evidence of somatic hypermutation. Importantly, the different identified antibody clones had diverse heavy chain CDR3 and deduced binding surfaces as suggested by structure modeling. This may suggest a potentially great heterogeneity in human antibodies recognizing PC-related epitopes on apoptotic cells. To re-construct the postulated structural format of the parental anti-PC antibody, the dominant clone was also expressed as a recombinant human polymeric IgM, which revealed a substantially increased binding reactivity, with dose-dependent and antigen-inhibitable binding of apoptotic cells. Our findings may have implication for improved prognostic testing and therapeutic interventions in human inflammatory disease.

  17. Detection of antibodies to co-trimoxazole (preservative drug interfering with routine red cell antibody screening

    Directory of Open Access Journals (Sweden)

    Deepti Sachan

    2018-01-01

    Full Text Available Drug-dependent antibodies can rarely cause interference in pretransfusion antibody screening. The diluents for commercial reagent red blood cells contain different antibiotics, such as chloramphenicol, neomycin sulfate, and gentamycin as a preservative. The presence of antibodies to a given drug in patient may lead to positive results when performing antibody identification. We present a rare case of detection of anti-co-trimoxazole antibody during routine antibody screening in a female patient undergoing neurosurgery. These antibodies mimicked as antibody against high-frequency red cell antigens reacting in both saline phase as well as antiglobulin phase. Anti-co-trimoxazole antibody was confirmed by repeating antibody screen using reagent red cells of different manufacturers with and without co-trimoxazole drug as preservative as well as using washed red cell panels. There were no associated clinical or laboratory evidence of hemolysis.

  18. Characterization of a monoclonal antibody with specificity for holo-transcobalamin

    Directory of Open Access Journals (Sweden)

    Fedosov Sergey N

    2006-01-01

    Full Text Available Abstract Background Holotranscobalamin, cobalamin-saturated transcobalamin, is the minor fraction of circulating cobalamin (vitamin B12, which is available for cellular uptake and hence is physiologically relevant. Currently, no method allows simple, direct quantification of holotranscobalamin. We now report on the identification and characterization of a monoclonal antibody with a unique specificity for holotranscobalamin. Methods The specificity and affinity of the monoclonal antibodies were determined using surface plasmon resonance and recombinant transcobalamin as well as by immobilizing the antibodies on magnetic microspheres and using native transcobalamin in serum. The epitope of the holotranscobalamin specific antibody was identified using phage display and comparison to a de novo generated three-dimensional model of transcobalamin using the program Rosetta. A direct assay for holotrnscobalamin in the ELISA format was developed using the specific antibody and compared to the commercial assay HoloTC RIA. Results An antibody exhibiting >100-fold specificity for holotranscobalamin over apotranscobalamin was identified. The affinity but not the specificity varied inversely with ionic strength and pH, indicating importance of electrostatic interactions. The epitope was discontinuous and epitope mapping of the antibody by phage display identified two similar motifs with no direct sequence similarity to transcobalamin. A comparison of the motifs with a de novo generated three-dimensional model of transcobalamin identified two structures in the N-terminal part of transcobalamin that resembled the motif. Using this antibody an ELISA based prototype assay was developed and compared to the only available commercial assay for measuring holotranscobalamin, HoloTC RIA. Conclusion The identified antibody possesses a unique specificity for holotranscobalamin and can be used to develop a direct assay for the quantification of holotranscobalamin.

  19. Solid phase double-antibody radioimmunoassay procedure

    International Nuclear Information System (INIS)

    Niswender, G.D.

    1977-01-01

    The present invention is concerned with the radioimmunoassay (RIA) procedure for assaying body fluid content of an antigenic substance which may either be an antigen itself or a hapten capable of being converted, such as by means of reaction with a protein, to an antigenic material. The present invention is concerned with a novel and improved modification of a double-antibody RIA technique in which there is a first antibody that is specific to the antigenic substance suspected to be present in a body fluid from which the assay is intended. The second antibody, however, is not specific to the antigenic substance or analyte, but is an antibody against the first antibody

  20. Production of Monoclonal Antibody against Human Nestin

    OpenAIRE

    Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad Mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

    2010-01-01

    We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140?250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such a...

  1. Antibody pretargeting advances cancer radioimmunodetection and radioimmunotherapy.

    Science.gov (United States)

    Goldenberg, David M; Sharkey, Robert M; Paganelli, Giovanni; Barbet, Jacques; Chatal, Jean-François

    2006-02-10

    This article reviews the methods of pretargeting, which involve separating the targeting antibody from the subsequent delivery of an imaging or therapeutic agent that binds to the tumor-localized antibody. This provides enhanced tumor:background ratios and the delivery of a higher therapeutic dose than when antibodies are directly conjugated with radionuclides, as currently practiced in cancer radioimmunotherapy. We describe initial promising clinical results using streptavidin-antibody constructs with biotin-radionuclide conjugates in the treatment of patients with malignant gliomas, and of bispecific antibodies with hapten-radionuclides in the therapy of tumors expressing carcinoembryonic antigen, such as medullary thyroid and small-cell lung cancers.

  2. Antibody Repertoire Development in Swine

    Czech Academy of Sciences Publication Activity Database

    Butler, J. E.; Wertz, N.; Šinkora, Marek

    2017-01-01

    Roč. 5, FEB 17 (2017), s. 255-279 ISSN 2165-8102 R&D Projects: GA ČR GA15-02274S; GA ČR(CZ) GA16-09296S Institutional support: RVO:61388971 Keywords : swine * pre-immune antibody repertoire * ileal Peyer's patches Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 4.708, year: 2016

  3. Memory B cell antibodies to HIV-1 gp140 cloned from individuals infected with clade A and B viruses.

    Directory of Open Access Journals (Sweden)

    Hugo Mouquet

    Full Text Available Understanding the antibody response to HIV-1 in humans that show broad neutralizing serologic activity is a crucial step in trying to reproduce such responses by vaccination. Investigating antibodies with cross clade reactivity is particularly important as these antibodies may target conserved epitopes on the HIV envelope gp160 protein. To this end we have used a clade B YU-2 gp140 trimeric antigen and single-cell antibody cloning methods to obtain 189 new anti-gp140 antibodies representing 51 independent B cell clones from the IgG memory B cells of 3 patients infected with HIV-1 clade A or B viruses and exhibiting broad neutralizing serologic activity. Our results support previous findings showing a diverse antibody response to HIV gp140 envelope protein, characterized by differentially expanded B-cell clones producing highly hypermutated antibodies with heterogenous gp140-specificity and neutralizing activity. In addition to their high-affinity binding to the HIV spike, the vast majority of the new anti-gp140 antibodies are also polyreactive. Although none of the new antibodies are as broad or potent as VRC01 or PG9, two clonally-related antibodies isolated from a clade A HIV-1 infected donor, directed against the gp120 variable loop 3, rank in the top 5% of the neutralizers identified in our large collection of 185 unique gp140-specific antibodies in terms of breadth and potency.

  4. Latency of Herpes Simplex Virus in Absence of Neutralizing Antibody: Model for Reactivation

    Science.gov (United States)

    Sekizawa, Tsuyoshi; Openshaw, Harry; Wohlenberg, Charles; Notkins, Abner Louis

    1980-11-01

    Mice inoculated with herpes simplex virus (type 1) by the lip or corneal route and then passively immunized with rabbit antibody to herpes simplex virus developed a latent infection in the trigeminal ganglia within 96 hours. Neutralizing antibody to herpes simplex virus was cleared from the circulation and could not be detected in most of these mice after 2 months. Examination of ganglia from the antibody-negative mice revealed latent virus in over 90 percent of the animals, indicating that serum neutralizing antibody is not necessary to maintain the latent state. When the lips or corneas of these mice were traumatized, viral reactivation occurred in up to 90 percent of the mice, as demonstrated by the appearance of neutralizing antibody. This study provides a model for identifying factors that trigger viral reactivation.

  5. Association analysis of anti-Epstein-Barr nuclear antigen-1 antibodies, anti-cyclic citrullinated peptide antibodies, the shared epitope and smoking status in Brazilian patients with rheumatoid arthritis

    Directory of Open Access Journals (Sweden)

    Michel Alexandre Yazbek

    2011-01-01

    Full Text Available INTRODUCTION: Epstein-Barr virus exposure appears to be an environmental trigger for rheumatoid arthritis that interacts with other risk factors. Relationships among anti-cyclic citrullinated peptide antibodies, the shared epitope, and smoking status have been observed in patients with rheumatoid arthritis from different populations. OBJECTIVE: To perform an association analysis of anti-Epstein-Barr nuclear antigen-1 antibodies, anti-cyclic citrullinated peptide antibodies, the shared epitope, and smoking status in Brazilian patients with rheumatoid arthritis. METHODS: In a case-control study, 140 rheumatoid arthritis patients and 143 healthy volunteers who were matched for age, sex, and ethnicity were recruited. Anti-Epstein-Barr nuclear antigen-1 antibodies and anti-cyclic citrullinated peptide antibodies were examined using an enzyme-linked immunosorbent assay, and shared epitope alleles were identified by genotyping. Smoking information was collected from all subjects. A comparative analysis of anti-Epstein-Barr nuclear antigen-1 antibodies, anti-cyclic citrullinated peptide antibodies, the shared epitope, and smoking status was performed in the patient group. Logistic regression analysis models were used to analyze the risk of rheumatoid arthritis. RESULTS: Anti-Epstein-Barr nuclear antigen-1 antibodies were not associated with anti-cyclic citrullinated peptide antibodies, shared epitope alleles, or smoking status. Anti-cyclic citrullinated peptide antibody positivity was significantly higher in smoking patients with shared epitope alleles (OR = 3.82. In a multivariate logistic regression analysis using stepwise selection, only anti-cyclic citrullinated peptide antibodies were found to be independently associated with rheumatoid arthritis (OR = 247.9. CONCLUSION: Anti-Epstein-Barr nuclear antigen-1 antibodies did not increase the risk of rheumatoid arthritis and were not associated with the rheumatoid arthritis risk factors studied. Smoking

  6. Binding induced conformational changes of proteins correlate with their intrinsic fluctuations: a case study of antibodies

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    Keskin Ozlem

    2007-05-01

    Full Text Available Abstract Background How antibodies recognize and bind to antigens can not be totally explained by rigid shape and electrostatic complimentarity models. Alternatively, pre-existing equilibrium hypothesis states that the native state of an antibody is not defined by a single rigid conformation but instead with an ensemble of similar conformations that co-exist at equilibrium. Antigens bind to one of the preferred conformations making this conformation more abundant shifting the equilibrium. Results Here, two antibodies, a germline antibody of 36–65 Fab and a monoclonal antibody, SPE7 are studied in detail to elucidate the mechanism of antibody-antigen recognition and to understand how a single antibody recognizes different antigens. An elastic network model, Anisotropic Network Model (ANM is used in the calculations. Pre-existing equilibrium is not restricted to apply to antibodies. Intrinsic fluctuations of eight proteins, from different classes of proteins, such as enzymes, binding and transport proteins are investigated to test the suitability of the method. The intrinsic fluctuations are compared with the experimentally observed ligand induced conformational changes of these proteins. The results show that the intrinsic fluctuations obtained by theoretical methods correlate with structural changes observed when a ligand is bound to the protein. The decomposition of the total fluctuations serves to identify the different individual modes of motion, ranging from the most cooperative ones involving the overall structure, to the most localized ones. Conclusion Results suggest that the pre-equilibrium concept holds for antibodies and the promiscuity of antibodies can also be explained this hypothesis: a limited number of conformational states driven by intrinsic motions of an antibody might be adequate to bind to different antigens.

  7. Auto-antibodies and their association with clinical findings in women diagnosed with microscopic colitis.

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    Bodil Roth

    Full Text Available BACKGROUND: Microscopic colitis (MC is a disease manifested by diarrhoea and is divided into collagenous and lymphocytic colitis. The aetiology is unknown, but auto-immunity is suggested. Auto-antibodies have been only rarely examined in this entity. The aim of the study was to examine the prevalence of auto-antibodies, and to examine associations between the presence of antibodies and clinical findings. METHODS AND FINDINGS: Women with MC verified by biopsy and younger than 73 years, at any Department of Gastroenterology, in the district of Skåne, between 2002 and 2010 were invited to participate in this study. The patients were asked to complete both a questionnaire describing their medical history and the Gastrointestinal Symptom Rating Scale (GSRS. Blood samples were collected. Anti-nuclear antibodies (ANA, anti-neutrophil cytoplasmic antibodies (ANCA, anti-Saccharomyces cerevisiae antibodies (ASCA, and antibodies against glutamic acid decarboxylase (anti-GAD, islet antigens-like insulin 2 (anti-IA2, thyroid peroxidase (anti-TPO, and thyrotropin receptor (TRAK were analysed. Of 240 women identified, 133 were finally included in the study, median age 63 (59-67 years. Apart from the MC diagnosis, 52% also suffered from irritable bowel syndrome, 31% from hypertension and 31% from allergy. The prevalence of ANA (14%, ASCA IgG (13%, and anti-TPO antibodies (14% for these patients was slightly higher than for the general population, and were found together with other concomitant diseases. Patients had more of all gastrointestinal symptoms compared with norm values, irrespective of antibody expression. CONCLUSIONS: Women with MC have a slightly increased prevalence of some auto-antibodies. These antibodies are not associated with symptoms, but are expressed in patients with concomitant diseases, obscuring the pathophysiology and clinical picture of MC.

  8. Expression of Human Immunodeficiency Virus Type 1 Neutralizing Antibody Fragments Using Human Vaginal Lactobacillus.

    Science.gov (United States)

    Marcobal, Angela; Liu, Xiaowen; Zhang, Wenlei; Dimitrov, Antony S; Jia, Letong; Lee, Peter P; Fouts, Timothy R; Parks, Thomas P; Lagenaur, Laurel A

    Eradication of human immunodeficiency virus type 1 (HIV-1) by vaccination with epitopes that produce broadly neutralizing antibodies is the ultimate goal for HIV prevention. However, generating appropriate immune responses has proven difficult. Expression of broadly neutralizing antibodies by vaginal colonizing lactobacilli provides an approach to passively target these antibodies to the mucosa. We tested the feasibility of expressing single-chain and single-domain antibodies (dAbs) in Lactobacillus to be used as a topical microbicide/live biotherapeutic. Lactobacilli provide an excellent platform to express anti-HIV proteins. Broadly neutralizing antibodies have been identified against epitopes on the HIV-1 envelope and have been made into active antibody fragments. We tested single-chain variable fragment m9 and dAb-m36 and its derivative m36.4 as prototype antibodies. We cloned and expressed the antibody fragments m9, m36, and m36.4 in Lactobacillus jensenii-1153 and tested the expression levels and functionality. We made a recombinant L. jensenii 1153-1128 that expresses dAb-m36.4. All antibody fragments m9, m36, and m36.4 were expressed by lactobacilli. However, we noted the smaller m36/m36.4 were expressed to higher levels, ≥3 μg/ml. All L. jensenii-expressed antibody fragments bound to gp120/CD4 complex; Lactobacillus-produced m36.4 inhibited HIV-1 BaL in a neutralization assay. Using a TZM-bl assay, we characterized the breadth of neutralization of the m36.4. Delivery of dAbs by Lactobacillus could provide passive transfer of these antibodies to the mucosa and longevity at the site of HIV-1 transmission.

  9. Donor-derived HLA antibody production in patients undergoing SCT from HLA antibody-positive donors.

    Science.gov (United States)

    Taniguchi, K; Yoshihara, S; Maruya, E; Ikegame, K; Kaida, K; Hayashi, K; Kato, R; Inoue, T; Fujioka, T; Tamaki, H; Okada, M; Onuma, T; Fujii, N; Kusunoki, Y; Soma, T; Saji, H; Ogawa, H

    2012-10-01

    Pre-existing donor-specific HLA antibodies in patients undergoing HLA-mismatched SCT have increasingly been recognized as a risk factor for primary graft failure. However, the clinical implications of the presence of HLA antibodies in donors remain unknown. We prospectively examined 123 related donors for the presence of HLA antibodies by using a Luminex-based single antigen assay. Of these, 1/57 (1.8%) male, 6/27 (22%) parous female and 0/39 (0%) nonparous female donors were HLA antibody-positive. Then, we determined the presence of HLA antibodies in seven patients who received SCT from antibody-positive donors. Of these, four became HLA antibody-positive after SCT. The specificities of the antibodies that emerged in the patients closely resembled those of the antibodies found in the donors, indicating their production by donor-derived plasma cells. Moreover, the kinetics of the HLA antibody levels were similar in all four patients: levels started increasing within 1 week after SCT and peaked at days 10-21, followed by a gradual decrease. These results suggest that donor-derived HLA antibody production frequently occurs in patients undergoing SCT from antibody-positive donors. Further studies are warranted for clarifying the clinical significance of donor-derived HLA antibodies, including the role of these antibodies in post transplant platelet transfusion refractoriness.

  10. Construction of Rabbit Immune Antibody Libraries.

    Science.gov (United States)

    Nguyen, Thi Thu Ha; Lee, Jong Seo; Shim, Hyunbo

    2018-01-01

    Rabbits have distinct advantages over mice as a source of target-specific antibodies. They produce higher affinity antibodies than mice, and may elicit strong immune response against antigens or epitopes that are poorly immunogenic or tolerated in mice. However, a great majority of currently available monoclonal antibodies are of murine origin because of the wider availability of murine fusion partner cell lines and well-established tools and protocols for fusion and cloning of mouse hybridoma. Phage-display selection of antibody libraries is an alternative method to hybridoma technology for the generation of target-specific monoclonal antibodies. High-affinity monoclonal antibodies from nonmurine species can readily be obtained by constructing immune antibody libraries from B cells of the immunized animal and screening the library by phage display. In this article, we describe the construction of a rabbit immune Fab library for the facile isolation of rabbit monoclonal antibodies. After immunization, B-cell cDNA is obtained from the spleen of the animal, from which antibody variable domain repertoires are amplified and assembled into a Fab repertoire by PCR. The Fab genes are then cloned into a phagemid vector and transformed to E. coli, from which a phage-displayed immune Fab library is rescued. Such a library can be biopanned against the immunization antigen for rapid identification of high-affinity, target-specific rabbit monoclonal antibodies.

  11. Dual-Color Bioluminescent Sensor Proteins for Therapeutic Drug Monitoring of Antitumor Antibodies.

    Science.gov (United States)

    van Rosmalen, Martijn; Ni, Yan; Vervoort, Daan F M; Arts, Remco; Ludwig, Susann K J; Merkx, Maarten

    2018-03-06

    Monitoring the levels of therapeutic antibodies in individual patients would allow patient-specific dose optimization, with the potential for major therapeutic and financial benefits. Our group recently developed a new platform of bioluminescent sensor proteins (LUMABS; LUMinescent AntiBody Sensor) that allow antibody detection directly in blood plasma. In this study, we targeted four clinically important therapeutic antibodies, the Her2-receptor targeting trastuzumab, the anti-CD20 antibodies rituximab and obinutuzumab, and the EGFR-blocking cetuximab. A strong correlation was found between the affinity of the antibody binding peptide and sensor performance. LUMABS sensors with physiologically relevant affinities and decent sensor responses were obtained for trastuzumab and cetuximab using mimotope and meditope peptides, respectively, with affinities in the 10 -7 M range. The lower affinity of the CD20-derived cyclic peptide employed in the anti-CD20 LUMABS sensor ( K d = 10 -5 M), translated in a LUMABS sensor with a strongly attenuated sensor response. The trastuzumab and cetuximab sensors were further characterized with respect to binding kinetics and their performance in undiluted blood plasma. For both antibodies, LUMABS-based detection directly in plasma compared well to the analytical performance of commercial ELISA kits. Besides identifying important design parameters for the development of new LUMABS sensors, this work demonstrates the potential of the LUMABS platform for point-of-care detection of therapeutic antibodies.

  12. [Irregular antierythocytic antibodies outside of the ABO system in the perinatal period].

    Science.gov (United States)

    Baptista-González, H A; Rosenfeld-Mann, F; Pérez-Pérez, J D; Quintanar-García, E

    1991-11-01

    Included in this study are the results of the tests done with irregular anti-erythrocytic antibodies outside of the ABO system of women in reproductive stage. In 2,623 cases considered, 279 samples positive for the antibodies were found (10.6%). In 184 cases (65.9%), the antibodies were classified as "immune irregular". The most frequently found antibodies from this group were the anti-D (63.08%), followed by the anti-c (1.07%), the anti-Kell (0.72%), anti-C, anti-E and an anti-Rh system (0.36% for each one). In 52 cases (18.6%) there were irregular "natural" antibodies, predominantly the anti-Lewis (9.68%), followed by anti-P (5.02%), the anti-I (3.2%), an anti-M and an anti-N (0.36% each one). In 43 (15.4%) cases, we were not able to identify the specificity of the antibody found. We include a discussion and a comparison of the frequency of these antibodies in our population. Based on these data, we recommend the clinician to consider the existence and specificity of the irregular anti-erythrocytic antibodies in their obstetric patients, candidates to transfusional therapy, as well as those newborn with hemolytic diseases.

  13. Alloimmunization due to red cell antibodies in Rhesus positive Omani Pregnant Women: Maternal and Perinatal outcome

    Directory of Open Access Journals (Sweden)

    Tamima Al-Dughaishi

    2015-01-01

    Full Text Available Objective: This study is aimed to determine the prevalence of alloimmunization due to antibodies to red blood cell (RBC antigens (other than rhesus [Rh] antigen and report the maternal, perinatal, and neonatal outcomes. Materials and Methods: A retrospective review of medical records of all patients with minor RBCs antibodies alloimmunization who were followed and delivered at Sultan Qaboos University Hospital, Oman from June 2011 to June 2013. Maternal characteristics, antibody type, antibody titer in addition to perinatal and neonatal outcomes were reviewed. Results: There were 1160 patients with Rh positive status in the study. The most common ABO blood group was O, followed by A, B, and AB. We found 33 out of 1160 Rh positive women alloimmunized with minor RBCs antibodies that gave a prevalence of minor RBCs alloimmunization of 2.7%. The most frequent antibody was anti-E 38%, followed by anti-c 17% and anti-kell 17%. 6 of these 33 patients were identified to have significant antibody titer, and two cases showed evidence of fetal anemia. Only one case required an intrauterine blood transfusion. The most common neonatal complication was jaundice in 53%, followed by respiratory distress syndrome in 28%. Two cases complicated by neonatal anemia required a postnatal blood transfusion. Conclusion: Alloimmunization with anti-E, anti-c, and anti-kell were the most common antibodies among the study group. Minor RBCs alloimmunization was an important cause of neonatal morbidity.

  14. Potent Human Monoclonal Antibodies against SARS CoV, Nipah and Hendra Viruses

    Science.gov (United States)

    Prabakaran, Ponraj; Zhongyu, Zhu; Xiao, Xiaodong; Biragyn, Arya; Dimitrov, Antony S.; Broder, Christopher C.; Dimitrov, Dimiter S.

    2009-01-01

    Polyclonal antibodies have a century-old history of being effective against some viruses; recently, monoclonal antibodies (mAbs) have also shown success. The humanized mAb Synagis (palivizumab) remains still the only mAb against respiratory syncytial virus (RSV) infections approved by the U.S. Food and Drug Administration (FDA). Recently, several potent human monoclonal antibodies (hmAbs) targeting the Severe Acute Respiratory Syndrome-Associated coronavirus (SARS CoV) S glycoproteins were developed quickly after the virus was identified in 2003. Among these antibodies, m396 and S230.15 exhibit exceptional potency and cross-reactivity as they neutralize isolates from the first and second outbreaks and from palm civets both in vitroand in mice. Similarly, the first fully hmAbs against two other paramyxoviruses, Hendra virus (HeV) and Nipah virus (NiV), which can cause up to 75% mortality, were recently developed; one of them, m102.4, shows exceptional cross-reactive potency against both NiV and HeV. Three-dimensional molecular structures of envelope glycoproteins from these viruses in complexes with antibodies and/or receptors were recently determined. Structural analyses along with other experiments have provided insights into the molecular mechanisms of receptor recognition and antibody neutralization, and suggested that these antibodies alone or in combination could successfully fight the viruses’ heterogeneity and mutability which is a major problem in the development of effective therapeutic agents against viruses, including therapeutic antibodies. PMID:19216624

  15. Characterization of rat basophilic leukemia cell surface proteins using monoclonal antibodies

    International Nuclear Information System (INIS)

    Buonocore-Buzzelli, L.M.

    1988-01-01

    Rat basophilic leukemia (RBL) cells express both immunoglobulin E (IgE) and immunoglobulin G (IgG) receptors. In this study, mouse monoclonal antibodies were produced against the RBL cell and screened for their ability to precipitate specific bands from 125 I surface labeled cells. Fourteen hybridomas were selected and divided into five groups since many of the hybridomas precipitated bands of identical molecular weight. One or more of the hybridomas from each group, and the cell surface antigens they identified, were further characterized. Binding of all the monoclonal antibodies to the RBL-2H3 cell surface was saturable and of high affinity. In cross inhibition studies, two of the antibodies were found to bind to identical or neighboring epitopes, presumably on the same cell surface molecule. Binding studies using other cell populations demonstrated that the monoclonal antibodies react not only with commonly expressed rat cell surface molecules but also with molecules specifically expressed on rat mast cells and basophils. None of the antibodies were found to induce or inhibit serotonin release from the RBL cells. Western blotting showed most of the antibodies to react with bands whose molecular weights resembled those seen by immuno-precipitation. Antibodies number sign 8 and number sign 12, although from the same group, were found to react with different subunits of the same cell surface protein. Sequential immunoprecipitation and peptide mapping confirmed that the antigens defined by these antibodies were structurally related

  16. Early developability screen of therapeutic antibody candidates using Taylor dispersion analysis and UV area imaging detection.

    Science.gov (United States)

    Lavoisier, Alexandra; Schlaeppi, Jean-Marc

    2015-01-01

    Therapeutic antibodies represent one of the fastest growing segments in the pharmaceutical market. They are used in a broad range of disease fields, such as autoimmune diseases, cancer, inflammation and infectious diseases. The growth of the segment has necessitated development of new analytical platforms for faster and better antibody selection and characterization. Early quality control and risk assessment of biophysical parameters help prevent failure in later stages of antibody development, and thus can reduce costs and save time. Critical parameters such as aggregation, conformational stability, colloidal stability and hydrophilicity, are measured during the early phase of antibody generation and guide the selection process of the best lead candidates in terms of technical developability. We report on the use of a novel instrument (ActiPix/Viscosizer) for measuring both the hydrodynamic radius and the absolute viscosity of antibodies based on Taylor dispersion analysis and UV area imaging. The looped microcapillary-based method combines low sample consumption, fast throughput and high precision compared to other conventional methods. From a random panel of 130 antibodies in the early selection process, we identified some with large hydrodynamic radius outside the normal distribution and others with non-Gaussian Taylor dispersion profiles. The antibodies with such abnormal properties were confirmed later in the selection process to show poor developability profiles. Moreover, combining these results with those of the viscosity measurements at high antibody concentrations allows screening, with limited amounts of materials, candidates with potential issues in pre-formulation development.

  17. Monoclonal antibodies that bind the renal Na/sup +//glucose symport system. 1. Identification

    Energy Technology Data Exchange (ETDEWEB)

    Wu, J.S.R.; Lever, J.E.

    1987-09-08

    Phlorizin is a specific, high-affinity ligand that binds the active site of the Na/sup +//glucose symporter by a Na/sup +/-dependent mechanism but is not itself transported across the membrane. The authors have isolated a panel of monoclonal antibodies that influence high-affinity, Na/sup +/-dependent phlorizin binding to pig renal brush border membranes. Antibodies were derived after immunization of mice either with highly purified renal brush border membranes or with apical membranes purified from LLC-PK/sub 1/, a cell line of pig renal proximal tubule origin. Antibody 11A3D6, an IgG/sub 2b/, reproducibly stimulated Na/sup +/-dependent phlorizin binding whereas antibody 18H10B12, an IgM, strongly inhibited specific binding. These effects were maximal after 30-min incubation and exhibited saturation at increased antibody concentrations. Antibodies did not affect Na/sup +/-dependent sugar uptake in vesicles but significantly prevented transport inhibition by bound phlorizin. Antibodies recognized a 75-kDa antigen identified by Western blot analysis of brush border membranes, and a 75-kDa membrane protein could be immunoprecipitated by 18H10B12. These properties, provide compelling evidence that the 75-kDa antigen recognized by these antibodies is a component of the renal Na/sup +//glucose symporter.

  18. Thrombosis and antiphospholipid antibody syndrome during acute Q fever

    Science.gov (United States)

    Million, Matthieu; Bardin, Nathalie; Bessis, Simon; Nouiakh, Nadia; Douliery, Charlaine; Edouard, Sophie; Angelakis, Emmanouil; Bosseray, Annick; Epaulard, Olivier; Branger, Stéphanie; Chaudier, Bernard; Blanc-Laserre, Karine; Ferreira-Maldent, Nicole; Demonchy, Elisa; Roblot, France; Reynes, Jacques; Djossou, Felix; Protopopescu, Camelia; Carrieri, Patrizia; Camoin-Jau, Laurence; Mege, Jean-Louis; Raoult, Didier

    2017-01-01

    Abstract Q fever is a neglected and potentially fatal disease. During acute Q fever, antiphospholipid antibodies are very prevalent and have been associated with fever, thrombocytopenia, acquired heart valve disease, and progression to chronic endocarditis. However, thrombosis, the main clinical criterion of the 2006 updated classification of the antiphospholipid syndrome, has not been assessed in this context. To test whether thrombosis is associated with antiphospholipid antibodies and whether the criteria for antiphospholipid syndrome can be met in patients with acute Q fever, we conducted a cross-sectional study at the French National Referral Center for Q fever. Patients included were diagnosed with acute Q fever in our Center between January 2007 and December 2015. Each patient's history and clinical characteristics were recorded with a standardized questionnaire. Predictive factors associated with thrombosis were assessed using a rare events logistic regression model. IgG anticardiolipin antibodies (IgG aCL) assessed by an enzyme-linked immunosorbent assay were tested on the Q fever diagnostic serum. A dose-dependent relationship between IgG aCL levels and thrombosis was tested using a receiver operating characteristic (ROC) analysis. Of the 664 patients identified for inclusion in the study, 313 (47.1%) had positive IgG aCL and 13 (1.9%) were diagnosed with thrombosis. Three patients fulfilled the antiphospholipid syndrome criteria. After multiple adjustments, only positive IgG aCL (relative risk, 14.46 [1.85–113.14], P = .011) were independently associated with thrombosis. ROC analysis identified a dose-dependent relationship between IgG aCL levels and occurrence of thrombosis (area under curve, 0.83, 95%CI [0.73–0.93], P antiphospholipid antibodies are associated with thrombosis, thrombocytopenia, and acquired valvular heart disease. Antiphospholipid antibodies should be systematically assessed in acute Q fever patients. Hydroxychloroquine

  19. Rethinking Modal Gender in the Context of the Universe of Tn-Types: Definitions and Mathematical Models for Tonicity and Phonicity

    Directory of Open Access Journals (Sweden)

    Marcus Alessi Bittencourt

    2016-07-01

    Full Text Available An indisputable cornerstone of the Western music tradition, the dialectic opposition between the major and minor grammatical modal genders has always been present in the imagination of musicians and music theorists for centuries. Such dialectics of opposition is especially important in the context of nineteenth-century harmonic dualism, with its ideas of tonicity and phonicity. These concepts serve as the main foundation for the way harmonic dualism conceives the major and minor worlds: two worlds with equivalent rights and properties, but with opposed polarities. This paper presents a redefinition of the terms tonicity and phonicity, translating those concepts to the context of post-tonal music theory. The terminologies of generatrix, tonicity, root, phonicity, vertex, and azimuth are explained in this paper, followed by propositions of mathematical models for those concepts, which spring from Richard Parncutt’s root-salience model for pitch-class sets. In order to demonstrate the possibilities of using modal gender as a criterion for the study and classification of the universe of Tn-types, we will present a taxonomy of the 351 transpositional set types, which comprises the categories of tonic (major, phonic (minor and neutral (genderless. In addition, there will be a small discussion on the effect of set symmetries and set asymmetries on the tonic/phonic properties of a Tn-type.

  20. TN trademark FLEX: a new generation of fluorocarbon o-rings developed by COGEMA logistics with enhanced characteristics at low temperature (-40 C)

    International Nuclear Information System (INIS)

    Issard, H.; Andre, R.

    2004-01-01

    Three main types of elastomers are used for the sealing of radioactive material transport casks with elastomeric gaskets: EPDM, fluorocarbons type Viton registered (standard designation: FKM) and silicon rubbers. Each rubber has specific characteristics in terms of temperature range, permeability, coefficient of expansion.. For the casks where high temperatures can be reached (200 C in continuous using), FKM gaskets are generally used. The problem is that this type of gasket does not guarantee the leaktightness at -40 C, which is a regulatory requirement. Two solutions are generally used: to specify a minimum heat load or a minimum ambient temperature. The direct consequence is that it is impossible to get B(U) approvals on the new concepts when FKM gaskets are used but only B(M) approvals, which generate significant additional justification costs (multiple submittals of Safety Analysis Reports, calculation of the minimum heat load or of the minimum ambient temperature..). Thus, it is important to develop gaskets with the same performance as FKM gaskets at high temperature but with enhanced performance at low temperature (and mainly, which guarantee the leaktightness at -40 C). COGEMA LOGISTICS has qualified a new generation of fluorocarbon O-rings (TN trademark FLEX gaskets) which can be used in continuous service on a -47 C/+200 C temperature range. TN trademark FLEX gaskets will be implemented on new casks designs

  1. Belimumab: anti-BLyS human monoclonal antibody, anti-BLyS monoclonal antibody, BmAb, human monoclonal antibody to B-lymphocyte stimulator.

    Science.gov (United States)

    2008-01-01

    Belimumab is a fully human monoclonal antibody that specifically recognizes and inhibits the biological activity of B-lymphocyte stimulator, or BLyS. Belimumab is in phase III trials for the treatment of systemic lupus erythematosus (SLE) and has completed a phase II trial in rheumatoid arthritis (RA); the product may also have potential in the treatment of other autoimmune disorders. In May 2001, Cambridge Antibody Technology (now MedImmune) completed its discovery programme and Human Genome Sciences identified belimumab as a candidate for clinical development. More than 1000 distinct human antibodies specific to BLyS were characterized by the collaboration.B-lymphocyte stimulator is a naturally occurring protein discovered by Human Genome Sciences that stimulates B-lymphocytes to develop into mature B cells. Laboratory studies have indicated that higher than normal levels of B-lymphocyte stimulator may contribute to the pathogenesis of autoimmune diseases, such as SLE and RA. Human Genome Sciences (HGS) and Cambridge Antibody Technology signed a collaborative agreement in August 1999 to study the B-lymphocyte stimulator as a human protein target. HGS is also developing other BLyS products. In March 2000, HGS and Cambridge Antibody Technology expanded their agreement into a 10-year collaboration and product development alliance, providing Human Genome Sciences with the right to use the antibody technology of Cambridge Antibody Technology to fully develop human antibodies for therapeutic and diagnostic purposes. Cambridge Antibody Technology will receive royalty payments on product sales from HGS, as well as the development and milestone payments it has already received. Belimumab will be manufactured in Human Genome Sciences' manufacturing facility, located in Rockville, MD, USA. HGS holds commercial rights to the drug. In July 2005, GlaxoSmithKline (GSK) exercised its co-development and co-promotion option to belimumab. In an agreement made in June 1996, HGS had

  2. Netrin-1 receptor antibodies in thymoma-associated neuromyotonia with myasthenia gravis.

    Science.gov (United States)

    Torres-Vega, Estefanía; Mancheño, Nuria; Cebrián-Silla, Arantxa; Herranz-Pérez, Vicente; Chumillas, María J; Moris, Germán; Joubert, Bastien; Honnorat, Jérôme; Sevilla, Teresa; Vílchez, Juan J; Dalmau, Josep; Graus, Francesc; García-Verdugo, José Manuel; Bataller, Luis

    2017-03-28

    To identify cell-surface antibodies in patients with neuromyotonia and to describe the main clinical implications. Sera of 3 patients with thymoma-associated neuromyotonia and myasthenia gravis were used to immunoprecipitate and characterize neuronal cell-surface antigens using reported techniques. The clinical significance of antibodies against precipitated proteins was assessed with sera of 98 patients (neuromyotonia 46, myasthenia gravis 52, thymoma 42; 33 of them with overlapping syndromes) and 219 controls (other neurologic diseases, cancer, and healthy volunteers). Immunoprecipitation studies identified 3 targets, including the Netrin-1 receptors DCC (deleted in colorectal carcinoma) and UNC5A (uncoordinated-5A) as well as Caspr2 (contactin-associated protein-like 2). Cell-based assays with these antigens showed that among the indicated patients, 9 had antibodies against Netrin-1 receptors (7 with additional Caspr2 antibodies) and 5 had isolated Caspr2 antibodies. Only one of the 219 controls had isolated Caspr2 antibodies with relapsing myelitis episodes. Among patients with neuromyotonia and/or myasthenia gravis, the presence of Netrin-1 receptor or Caspr2 antibodies predicted thymoma ( p thymoma, myasthenia gravis, and neuromyotonia, often with Morvan syndrome ( p = 0.009). Expression of DCC, UNC5A, and Caspr2 proteins was demonstrated in paraffin-embedded thymoma samples (3) and normal thymus. Antibodies against Netrin-1 receptors (DCC and UNC5a) and Caspr2 often coexist and associate with thymoma in patients with neuromyotonia and myasthenia gravis. This study provides Class III evidence that antibodies against Netrin-1 receptors can identify patients with thymoma (sensitivity 21.4%, specificity 100%). © 2017 American Academy of Neurology.

  3. Update on antiphospholipid antibody syndrome.

    Science.gov (United States)

    Lopes, Michelle Remião Ugolini; Danowski, Adriana; Funke, Andreas; Rêgo, Jozelia; Levy, Roger; Andrade, Danieli Castro Oliveira de

    2017-11-01

    Antiphospholipid syndrome (APS) is an autoimmune disease characterized by antiphospholipid antibodies (aPL) associated with thrombosis and/or pregnancy morbidity. Most APS events are directly related to thrombotic events, which may affect small, medium or large vessels. Other clinical features like thrombocytopenia, nephropathy, cardiac valve disease, cognitive dysfunction and skin ulcers (called non-criteria manifestations) add significant morbidity to this syndrome and represent clinical situations that are challenging. APS was initially described in patients with systemic lupus erythematosus (SLE) but it can occur in patients without any other autoimmune disease. Despite the autoimmune nature of this syndrome, APS treatment is still based on anticoagulation and antiplatelet therapy.

  4. Fully automated antibody structure prediction using BIOVIA tools: Validation study.

    Directory of Open Access Journals (Sweden)

    Helen Kemmish

    Full Text Available We describe the methodology and results from our validation study of the fully automated antibody structure prediction tool available in the BIOVIA (formerly Accelrys protein modeling suite. Extending our previous study, we have validated the automated approach using a larger and more diverse data set (157 unique antibody Fv domains versus 11 in the previous study. In the current study, we explore the effect of varying several parameter settings in order to better understand their influence on the resulting model quality. Specifically, we investigated the dependence on different methods of framework model construction, antibody numbering schemes (Chothia, IMGT, Honegger and Kabat, the influence of compatibility of loop templates using canonical type filtering, wider exploration of model solution space, and others. Our results show that our recently introduced Top5 framework modeling method results in a small but significant improvement in model quality whereas the effect of other parameters is not significant. Our analysis provides improved guidelines of best practices for using our protocol to build antibody structures. We also identify some limitations of the current computational model which will enhance proper evaluation of model quality by users and suggests possible future enhancements.

  5. Antinuclear antibodies as ancillary markers in primary biliary cirrhosis.

    Science.gov (United States)

    Granito, Alessandro; Muratori, Paolo; Quarneti, Chiara; Pappas, Georgios; Cicola, Ronny; Muratori, Luigi

    2012-01-01

    Antimitochondrial antibodies are the serological hallmark of primary biliary cirrhosis (PBC). Besides antimitochondrial antibodies, the autoantibody profile of PBC includes antinuclear antibodies (ANA) which are detectable by indirect immunofluorescence in up to 50% of PBC patients. Two immunofluorescence patterns are considered 'PBC-specific': the multiple nuclear dots and rim-like/membranous patterns. The target antigens of the multiple nuclear dots pattern have been identified as Sp100 and promyelocytic leukemia protein, whereas the rim-like/membranous pattern is given by autoantibodies recognizing multiple proteins such as gp210, nucleoporin p62 and the lamin B receptor. Other ANA, especially those already known in the rheumatological setting, such as anticentromere, anti-SSA/Ro and anti-dsDNA antibodies, can be frequently found in PBC, often coexisting in the same patient. In this article, we will report on recent progress in the antigenic characterization of ANA in PBC, their detection with both traditional assays and Western blot/ELISA with molecularly defined nuclear antigens, and we will discuss their clinical significance.

  6. Detection of antibodies to Helicobacter pylori cell surface antigens.

    Science.gov (United States)

    Guruge, J L; Schalén, C; Nilsson, I; Ljungh, A; Tyszkiewicz, T; Wikander, M; Wadström, T

    1990-01-01

    Serum IgG antibodies of Helicobacter pylori were detected in single-dilution ELISA using glycine extracted material. Among 148 endoscopy patients 59% displayed antibodies; as expected, a higher occurrence (90%) was found in patients with positive gastric culture for H. pylori than in culture negative patients (37%). Among 68 blood donors the frequency of H. pylori antibodies was 28%. In 73 children less than 15 years of age examined for unrelated disorders the occurrence was 4%. By immunoblotting using the same extract, 3 prominent bands, 29K, 54K and 60K and several weak bands were identified. These were formed by 57%, 92%, and 65%, respectively, of the ELISA positive patient sera. Comparing culture positive and negative patients, the 3 bands occurred more often among the culture positive subjects though between 18 and 61% of the sera from culture negative patients gave either of the bands. When comparing the glycine extracts of 4 different H. pylori strains with separate haemagglutinating patterns no differences in the position of the major bands emerged. By absorption experiments no immunological cross-reactivity with components of Escherichia coli, Klebsiella pneumoniae, Campylobacter jejuni or C. fetus was found. Thus, the glycine extract seemed specific for the detection of antibodies to H. pylori.

  7. Surveillance for Intracellular Antibody by Cytosolic Fc Receptor TRIM21

    Directory of Open Access Journals (Sweden)

    William A. McEwan

    2016-11-01

    Full Text Available TRIM21 has emerged as an atypical Fc receptor that is broadly conserved and widely expressed in the cytoplasm of mammalian cells. Viruses that traffic surface-bound antibodies into the cell during infection recruit TRIM21 via a high affinity interaction between Fc and TRIM21 PRYSPRY domain. Following binding of intracellular antibody, TRIM21 acts as both antiviral effector and sensor for innate immune signalling. These activities serve to reduce viral replication by orders of magnitude in vitro and contribute to host survival during in vivo infection. Neutralization occurs rapidly after detection and requires the activity of the ubiquitin-proteasome system. The microbial targets of this arm of intracellular immunity are still being identified: TRIM21 activity has been reported following infection by several non-enveloped viruses and intracellular bacteria. These findings extend the sphere of influence of antibodies to the intracellular domain and have broad implications for immunity. TRIM21 has been implicated in the chronic auto-immune condition systemic lupus erythematosus and is itself an auto-antigen in Sjögren’s syndrome. This review summarises our current understanding of TRIM21’s role as a cytosolic Fc receptor and briefly discusses pathological circumstances where intracellular antibodies have been described, or are hypothesized to occur, and may benefit from further investigations of the role of TRIM21.

  8. Identification, production, and use of polyol-responsive monoclonal antibodies for immunoaffinity chromatography.

    Science.gov (United States)

    Thompson, Nancy E; Foley, Katherine M; Stalder, Elizabeth S; Burgess, Richard R

    2009-01-01

    Immunoaffinity chromatography is a powerful tool for purification of proteins and protein complexes. The availability of monoclonal antibodies (mAbs) has revolutionized the field of immunoaffinity chromatography by providing a continuous supply of highly uniform antibody. Before the availability of mAbs, the recovery of the target protein from immobilized polyclonal antibodies usually required very harsh, often denaturing conditions. Although harsh conditions are often still used to disrupt the antigen-antibody interaction when using a mAb, various methods have been developed to exploit the uniformity of the antigen-antibody reaction in order to identify agents or conditions that gently disrupt this interaction and thus result in higher recovery of active protein from immunoaffinity chromatography. We discuss here the use of a specific type of monoclonal antibody that we have designated "polyol-responsive monoclonal antibodies" (PR-mAbs). These are naturally occurring mAbs that have high affinity for the antigen under binding conditions, but have low affinity in the presence of a combination of low molecular weight hydroxylated compounds (polyols) and nonchaotropic salts. Therefore, these PR-mAbs can be used for gentle immunoaffinity chromatography. PR-mAbs can be easily identified and adapted to a powerful protein purification method for a target protein.

  9. Antiphospholipid antibodies and non-thrombotic manifestations of systemic lupus erythematosus.

    Science.gov (United States)

    İlgen, U; Yayla, M E; Ateş, A; Okatan, İ E; Yurteri, E U; Torgutalp, M; Keleşoğlu, A B D; Turgay, T M; Kınıklı, G

    2018-04-01

    Objectives The aim of this study was to investigate the association between antiphospholipid antibodies and non-thrombotic and non-gestational manifestations of systemic lupus erythematosus. Methods Systemic lupus erythematosus patients with persistently positive antiphospholipid antibodies or lupus anticoagulant were identified and grouped as systemic lupus erythematosus with antiphospholipid syndrome (SLE-APS), systemic lupus erythematosus with positive antiphospholipid antibodies/lupus anticoagulant without antiphospholipid syndrome (SLE-aPL), and systemic lupus erythematosus with negative aPLs (SLE-No aPL). Groups were compared in terms of non-thrombotic systemic lupus erythematosus manifestations and laboratory features retrospectively. Results A total of 150 systemic lupus erythematosus patients, 26 with SLE-APS, 25 with SLE-aPL, and 99 with SLE-No aPL, were identified. Livedo reticularis, neurologic involvement, and thrombocytopenia were more common in antiphospholipid antibody positive systemic lupus erythematosus cases. Malar rash, arthritis, and pleuritis were more common in the SLE-No aPL, SLE-APS, and SLE-aPL groups, respectively. Positivity rates and titers of specific antiphospholipid antibodies did not differ between the SLE-APS and SLE-aPL groups. Conclusions Presence of antiphospholipid syndrome or persistent antiphospholipid antibodies may be related to non-thrombotic and non-gestational systemic lupus erythematosus manifestations. Patients with systemic lupus erythematosus plus antiphospholipid syndrome and persistent antiphospholipid antibodies without antiphospholipid syndrome also differ in terms of systemic lupus erythematosus manifestations.

  10. Phase Separation in Solutions of Monoclonal Antibodies

    Science.gov (United States)

    Benedek, George; Wang, Ying; Lomakin, Aleksey; Latypov, Ramil

    2012-02-01

    We report the observation of liquid-liquid phase separation (LLPS) in a solution of humanized monoclonal antibodies, IgG2, and the effects of human serum albumin, a major blood protein, on this phase separation. We find a significant reduction of phase separation temperature in the presence of albumin, and a preferential partitioning of the albumin into the antibody-rich phase. We provide a general thermodynamic analysis of the antibody-albumin mixture phase diagram and relate its features to the magnitude of the effective inter-protein interactions. Our analysis suggests that additives (HSA in this report), which have moderate attraction with antibody molecules, may be used to forestall undesirable protein condensation in antibody solutions. Our findings are relevant to understanding the stability of pharmaceutical solutions of antibodies and the mechanisms of cryoglobulinemia.

  11. The future of antibodies as cancer drugs.

    Science.gov (United States)

    Reichert, Janice M; Dhimolea, Eugen

    2012-09-01

    Targeted therapeutics such as monoclonal antibodies (mAbs) have proven successful as cancer drugs. To profile products that could be marketed in the future, we examined the current commercial clinical pipeline of mAb candidates for cancer. Our analysis revealed trends toward development of a variety of noncanonical mAbs, including antibody-drug conjugates (ADCs), bispecific antibodies, engineered antibodies and antibody fragments and/or domains. We found substantial diversity in the antibody sequence source, isotype, carbohydrate residues, targets and mechanisms of action (MOA). Although well-validated targets, such as epidermal growth factor receptor (EGFR) and CD20, continue to provide opportunities for companies, we found notable trends toward targeting less-well-validated antigens and exploration of innovative MOA such as the generation of anticancer immune responses or recruitment of cytotoxic T cells. Copyright © 2012 Elsevier Ltd. All rights reserved.

  12. Characterization of antibodies against ferret immunoglobulins, cytokines and CD markers

    DEFF Research Database (Denmark)

    Martel, Cyril Jean-Marie; Aasted, Bent

    2009-01-01

    Ferret IgG and IgM were purified from normal serum, while ferret IgA was purified from bile. The estimated molecular weights of the immunoglobulin gamma, alpha and mu heavy chains were found to be 54 kDa, 69 kDa and 83 kDa, respectively. For immunological (ELISA) quantification of ferret...... immunoglobulins, we identified and characterized polyclonal antibodies towards ferret IgG, IgM and IgA. We also identified 22 monoclonal antibodies (mAbs) raised mostly against human CD markers which cross-reacted with ferret leukocytes. These antibodies were originally specific against human CD8, CD9, CD14, CD18......, CD25, CD29, CD32, CD44, CD61, CD71, CD79b, CD88, CD104, CD172a and mink CD3. Finally, we identified 4 cross-reacting mAbs with specificities against ferret interferon-gamma, TNF-alpha, interleukin-4 and interleukin-8....

  13. Bilateral Internal Carotid Artery Occlusion Associated with the Antiphospholipid Antibody Syndrome

    Directory of Open Access Journals (Sweden)

    Pria Anand

    2014-03-01

    Full Text Available A 39-year-old woman presented with a right-hemispheric stroke 1 year after she had suffered a left-hemispheric stroke. Her diagnostic workup was notable for bilateral occlusions of the internal carotid arteries at their origins and a positive lupus anticoagulant antibody test. There was no evidence of carotid dissection or another identifiable cause for her carotid occlusions. These findings suggest that the antiphospholipid antibody syndrome may be implicated in the pathological changes that resulted in occlusions of the extracranial internal carotid arteries. Young stroke patients who present with unexplained internal carotid artery occlusions may benefit from testing for the presence of antiphospholipid antibodies.

  14. Microselection - Affinity Selecting Antibodies against a Single Rare Cell in a Heterogeneous Population

    DEFF Research Database (Denmark)

    Sørensen, Morten Dræby; Agerholm, Inge Errebo; Christensen, Britta

    2009-01-01

    antibodies. Here we have generated a microselection method allowing antibody selection, by phage display, targeting a single cell in a heterogeneous population. One K562 cell (female origin) was positioned on glass-slide among millions of lymphocytes from male donor, identifying the K562 cell by FISH (XX......). Several single cell selections were performed on such individual slides. The phage particles bound to the target cell is protected by a minute disc, while inactivating all remaining phage by UV-irradiation; leaving only the phage bound to the target cell viable. We hereby retrieved up to eight antibodies...

  15. Monoclonal Antibodies Against IL-13 and IL-31RA in Development for Atopic Dermatitis

    DEFF Research Database (Denmark)

    Hamann, Carsten R; Thyssen, Jacob P

    2017-01-01

    phase two trial has been completed for a humanized antibody against IL-31 receptor alpha, one subunit of the IL-31 receptor complex. This study showed significant dose-dependent reductions in pruritus, EASI scores, and markers of sleep quality. Initial clinical trials for monoclonal antibodies against......The IL-13 and IL-31 cytokines and inflammatory pathways have been identified as important for atopic dermatitis (AD) pathophysiology. Monoclonal antibodies against IL-13 have been studied for the treatment of asthma since 2011. More recently, two phase two trials have been completed...

  16. Prevalence of irregular red blood cell antibodies among healthy blood donors in Delhi population.

    Science.gov (United States)

    Garg, Neeraj; Sharma, Tanya; Singh, Bharat

    2014-06-01

    To evaluate the prevalence of the anti-red blood cell antibodies among healthy blood donors. Antibody screening of all voluntary blood donor serum was performed as routine immunohematological procedure. Positive sera were further investigated to identify the specificity of irregular erythrocyte antibody by commercially available red cell panel (ID-Dia Panel, Diamed-ID Microtyping System). A total of 47,450 donors were screened for the presence of irregular erythrocyte antibodies. A total of forty-six donors showed presence of alloantibodies in their serum (46/47,450%, 0.09%), yielding a prevalence of 0.09%. Most frequent alloantibodies identified were of MNS blood group system. The results showed statistically a higher prevalence of RBC alloantibodies in females than in males. Screening for presence of alloantibodies in donor blood is important to provide compatible blood products and to avoid transfusion reactions. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Warm antibody autoimmune hemolytic anemia.

    Science.gov (United States)

    Kalfa, Theodosia A

    2016-12-02

    Autoimmune hemolytic anemia (AIHA) is a rare and heterogeneous disease that affects 1 to 3/100 000 patients per year. AIHA caused by warm autoantibodies (w-AIHA), ie, antibodies that react with their antigens on the red blood cell optimally at 37°C, is the most common type, comprising ∼70% to 80% of all adult cases and ∼50% of pediatric cases. About half of the w-AIHA cases are called primary because no specific etiology can be found, whereas the rest are secondary to other recognizable underlying disorders. This review will focus on the postulated immunopathogenetic mechanisms in idiopathic and secondary w-AIHA and report on the rare cases of direct antiglobulin test-negative AIHA, which are even more likely to be fatal because of inherent characteristics of the causative antibodies, as well as because of delays in diagnosis and initiation of appropriate treatment. Then, the characteristics of w-AIHA associated with genetically defined immune dysregulation disorders and special considerations on its management will be discussed. Finally, the standard treatment options and newer therapeutic approaches for this chronic autoimmune blood disorder will be reviewed. © 2016 by The American Society of Hematology. All rights reserved.

  18. An anti vimentin antibody promotes tube formation

    DEFF Research Database (Denmark)

    Jørgensen, Mathias Lindh; Møller, Carina Kjeldahl; Rasmussen, Lasse

    2017-01-01

    antibody technology, promotes tube formation of endothelial cells in a 2D matrigel assay. By binding vimentin, the antibody increases the tube formation by 21% after 5 hours of incubation. Addition of the antibody directly to cultured endothelial cells does not influence endothelial cell migration...... or proliferation. The enhanced tube formation can be seen for up to 10 hours where after the effect decreases. It is shown that the antibody-binding site is located on the coil 2 domain of vimentin. To our knowledge this is the first study that demonstrates an enhanced tube formation by binding vimentin in a 2D...

  19. Uses of monoclonal antibody 8H9

    Energy Technology Data Exchange (ETDEWEB)

    Cheung, Nai-Kong V.

    2018-04-10

    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.

  20. Exceptional Antibodies Produced by Successive Immunizations.

    Directory of Open Access Journals (Sweden)

    Patricia J Gearhart

    2015-12-01

    Full Text Available Antibodies stand between us and pathogens. Viruses mutate quickly to avoid detection, and antibodies mutate at similar rates to hunt them down. This death spiral is fueled by specialized proteins and error-prone polymerases that change DNA sequences. Here, we explore how B lymphocytes stay in the race by expressing activation-induced deaminase, which unleashes a tsunami of mutations in the immunoglobulin loci. This produces random DNA substitutions, followed by selection for the highest affinity antibodies. We may be able to manipulate the process to produce better antibodies by expanding the repertoire of specific B cells through successive vaccinations.

  1. Affinity improvement of a therapeutic antibody by structure-based computational design: generation of electrostatic interactions in the transition state stabilizes the antibody-antigen complex.

    Directory of Open Access Journals (Sweden)

    Masato Kiyoshi

    Full Text Available The optimization of antibodies is a desirable goal towards the development of better therapeutic strategies. The antibody 11K2 was previously developed as a therapeutic tool for inflammatory diseases, and displays very high affinity (4.6 pM for its antigen the chemokine MCP-1 (monocyte chemo-attractant protein-1. We have employed a virtual library of mutations of 11K2 to identify antibody variants of potentially higher affinity, and to establish benchmarks in the engineering of a mature therapeutic antibody. The most promising candidates identified in the virtual screening were examined by surface plasmon resonance to validate the computational predictions, and to characterize their binding affinity and key thermodynamic properties in detail. Only mutations in the light-chain of the antibody are effective at enhancing its affinity for the antigen in vitro, suggesting that the interaction surface of the heavy-chain (dominated by the hot-spot residue Phe101 is not amenable to optimization. The single-mutation with the highest affinity is L-N31R (4.6-fold higher affinity than wild-type antibody. Importantly, all the single-mutations showing increase affinity incorporate a charged residue (Arg, Asp, or Glu. The characterization of the relevant thermodynamic parameters clarifies the energetic mechanism. Essentially, the formation of new electrostatic interactions early in the binding reaction coordinate (transition state or earlier benefits the durability of the antibody-antigen complex. The combination of in silico calculations and thermodynamic analysis is an effective strategy to improve the affinity of a matured therapeutic antibody.

  2. Antibodies with specificity for native and denatured forms of ovalbumin differ in reactivity between enzyme-linked immunosorbent assays

    DEFF Research Database (Denmark)

    Holm, B.E.; Bergmann, A.C.; Hansen, Paul Robert

    2015-01-01

    constituting amino acids 130–135 and 136–141, respectively. Moreover, comparison of antibody reactivity to N OVA revealed that in the streptavidin-capture ELISA, antibody reactivity was notably reduced compared to ELISA employing surface-bound OVA. Collectively, immunization with native OVA preferentially...... to native OVA reacted strongly with native and denatured OVA in both assays, but did not react with the overlapping peptides. Polyclonal antibodies to denatured OVA reacted strongly with both OVA forms and with several of the overlapping peptides. Monoclonal antibodies to native OVA reacted preferentially...... with three-dimensional epitopes on native OVA and not with denatured OVA. Monoclonal antibodies to denatured OVA showed reactivity to both OVA forms. Two of these monoclonal antibodies, HYB 94-06 and 94-07, showed reactivity to overlapping peptides and their epitopes were identified as flexible structures...

  3. High throughput discovery of influenza virus neutralizing antibodies from phage-displayed synthetic antibody libraries.

    Science.gov (United States)

    Chen, Ing-Chien; Chiu, Yi-Kai; Yu, Chung-Ming; Lee, Cheng-Chung; Tung, Chao-Ping; Tsou, Yueh-Liang; Huang, Yi-Jen; Lin, Chia-Lung; Chen, Hong-Sen; Wang, Andrew H-J; Yang, An-Suei

    2017-10-31

    Pandemic and epidemic outbreaks of influenza A virus (IAV) infection pose severe challenges to human society. Passive immunotherapy with recombinant neutralizing antibodies can potentially mitigate the threats of IAV infection. With a high throughput neutralizing antibody discovery platform, we produced artificial anti-hemagglutinin (HA) IAV-neutralizing IgGs from phage-displayed synthetic scFv libraries without necessitating prior memory of antibody-antigen interactions or relying on affinity maturation essential for in vivo immune systems to generate highly specific neutralizing antibodies. At least two thirds of the epitope groups of the artificial anti-HA antibodies resemble those of natural protective anti-HA antibodies, providing alternatives to neutralizing antibodies from natural antibody repertoires. With continuing advancement in designing and constructing synthetic scFv libraries, this technological platform is useful in mitigating not only the threats of IAV pandemics but also those from other newly emerging viral infections.

  4. Not All Antibodies Are Created Equal: Factors That Influence Antibody Mediated Rejection

    Directory of Open Access Journals (Sweden)

    Carrie L. Butler

    2017-01-01

    Full Text Available Consistent with Dr. Paul Terasaki’s “humoral theory of rejection” numerous studies have shown that HLA antibodies can cause acute and chronic antibody mediated rejection (AMR and decreased graft survival. New evidence also supports a role for antibodies to non-HLA antigens in AMR and allograft injury. Despite the remarkable efforts by leaders in the field who pioneered single antigen bead technology for detection of donor specific antibodies, a considerable amount of work is still needed to better define the antibody attributes that are associated with AMR pathology. This review highlights what is currently known about the clinical context of pre and posttransplant antibodies, antibody characteristics that influence AMR, and the paths after donor specific antibody production (no rejection, subclinical rejection, and clinical dysfunction with AMR.

  5. A generic mechanism in Neisseria meningitidis for enhanced resistance against bactericidal antibodies

    OpenAIRE

    Uria, Maria Jose; Zhang, Qian; Li, Yanwen; Chan, Angel; Exley, Rachel M.; Gollan, Bridget; Chan, Hannah; Feavers, Ian; Yarwood, Andy; Abad, Raquel; Borrow, Ray; Fleck, Roland A.; Mulloy, Barbara; Vazquez, Julio A.; Tang, Christoph M.

    2008-01-01

    The presence of serum bactericidal antibodies is a proven correlate of protection against systemic infection with the important human pathogen Neisseria meningitidis. We have identified three serogroup C N. meningitidis (MenC) isolates recovered from patients with invasive meningococcal disease that resist killing by bactericidal antibodies induced by the MenC conjugate vaccine. None of the patients had received the vaccine, which has been successfully introduced in countries in North America...

  6. Human anti-Dectin-1 antibody, hybridoma producing said antibody and applications thereof

    OpenAIRE

    Kremer, Leonor; Llorente Gómez, María de las Mercedes; Casasnovas, José María; Fernández Ruíz, Elena; Galán Díez, Marta

    2008-01-01

    [EN] The invention relates to hybridoma MGD3 and the monoclonal antibody produced thereby (also called MGD3), which specifically recognises the human Dectin-1 membrane receptor. Antibody MGD3 is capable of inhibiting the binding of Dectin-1 to the natural ligand thereof, the ss-glucans that are components of the fungal wall. In addition, the aforementioned antibody specifically blocks binding to Candida albicans and the secretion of cytokines induced thereby. The MGD3 antibody obtained enable...

  7. Stratification of Antibody-Positive Subjects by Antibody Level Reveals an Impact of Immunogenicity on Pharmacokinetics

    OpenAIRE

    Zhou, Lei; Hoofring, Sarah A.; Wu, Yu; Vu, Thuy; Ma, Peiming; Swanson, Steven J.; Chirmule, Narendra; Starcevic, Marta

    2012-01-01

    The availability of highly sensitive immunoassays enables the detection of antidrug antibody (ADA) responses of various concentrations and affinities. The analysis of the impact of antibody status on drug pharmacokinetics (PK) is confounded by the presence of low-affinity or low-concentration antibody responses within the dataset. In a phase 2 clinical trial, a large proportion of subjects (45%) developed ADA following weekly dosing with AMG 317, a fully human monoclonal antibody therapeutic....

  8. Positivity for antiribonucleoprotein antibodies in rheumatic diseases: focus on scleroderma systematica

    Directory of Open Access Journals (Sweden)

    R. U. Shayakhmetova

    2017-01-01

    Full Text Available The review deals with the problem of antibody production in scleroderma systematica (SDS, with a focus on antiribonucleoprotein (RNP antibodies: frequency, structure, and clinical associations. SDS is a progressive polysyndromic disease with characteristic lesion of the skin, locomotor apparatus, viscera (heart, lungs, digestive tract, and kidneys, and common vasospastic disorders as Raynaud's syndrome, the basis for which are the processes of connective tissue disorganization with a predominance of fibrosclerotic changes and vascular pathology as peculiar endarteritis obliterans. The presence of antibodies to different autoantigens is a distinguishing feature of SDS. Among the patients who meet the classification criteria for SDS, there is a subgroup of patients who are not found to have SDS-specific antinuclear antibodies, but have antibodies to soluble nuclear autoantigens, namely, antibodies to RNP. This type of autoantibodies is described in various systemic connective tissue diseases: systemic lupus erythematosus, dermatomyositis, polymyositis, and SDS. The detection rate of anti-RNP antibodies in SDS varies from 5 to 30% in different ethnic groups. In addition, positivity for anti-RNP antibodies is a characteristic feature of mixed connective tissue disease. A more detailed study of anti-U1-RNP antibody-positive patients with SDS, by identifying a new subtype of SDS, comparing, and searching for dissimilarities of anti-U1-RNP antibody carriers from other well-described phenotypes of SDS, is of interest today. This problem is relevant, by taking into account the personalized approach to following up patients, which is actively being elaborated in rheumatology. 

  9. RNA recognition by a human antibody against brain cytoplasmic 200 RNA.

    Science.gov (United States)

    Jung, Euihan; Lee, Jungmin; Hong, Hyo Jeong; Park, Insoo; Lee, Younghoon

    2014-06-01

    Diverse functional RNAs participate in a wide range of cellular processes. The RNA structure is critical for function, either on its own or as a complex form with proteins and other ligands. Therefore, analysis of the RNA conformation in cells is essential for understanding their functional mechanisms. However, no appropriate methods have been established as yet. Here, we developed an efficient strategy for panning and affinity maturation of anti-RNA human monoclonal antibodies from a naïve antigen binding fragment (Fab) combinatorial phage library. Brain cytoplasmic 200 (BC200) RNA, which is also highly expressed in some tumors, was used as an RNA antigen. We identified MabBC200-A3 as the optimal binding antibody. Mutagenesis and SELEX experiments showed that the antibody recognized a domain of BC200 in a structure- and sequence-dependent manner. Various breast cancer cell lines were further examined for BC200 RNA expression using conventional hybridization and immunoanalysis with MabBC200-A3 to see whether the antibody specifically recognizes BC200 RNA among the total purified RNAs. The amounts of antibody-recognizable BC200 RNA were consistent with hybridization signals among the cell lines. Furthermore, the antibody was able to discriminate BC200 RNA from other RNAs, supporting the utility of this antibody as a specific RNA structure-recognizing probe. Intriguingly, however, when permeabilized cells were subjected to immunoanalysis instead of purified total RNA, the amount of antibody-recognizable RNA was not correlated with the cellular level of BC200 RNA, indicating that BC200 RNA exists as two distinct forms (antibody-recognizable and nonrecognizable) in breast cancer cells and that their distribution depends on the cell type. Our results clearly demonstrate that anti-RNA antibodies provide an effective novel tool for detecting and analyzing RNA conformation. © 2014 Jung et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  10. Structure of a Human Astrovirus Capsid-Antibody Complex and Mechanistic Insights into Virus Neutralization

    Energy Technology Data Exchange (ETDEWEB)

    Bogdanoff, Walter A.; Campos, Jocelyn; Perez, Edmundo I.; Yin, Lu; Alexander, David L.; DuBois, Rebecca M. (UCSC)

    2016-11-02

    ABSTRACT

    Human astroviruses (HAstVs) are a leading cause of viral diarrhea in young children, the immunocompromised, and the elderly. There are no vaccines or antiviral therapies against HAstV disease. Several lines of evidence point to the presence of protective antibodies in healthy adults as a mechanism governing protection against reinfection by HAstV. However, development of anti-HAstV therapies is hampered by the gap in knowledge of protective antibody epitopes on the HAstV capsid surface. Here, we report the structure of the HAstV capsid spike domain bound to the neutralizing monoclonal antibody PL-2. The antibody uses all six complementarity-determining regions to bind to a quaternary epitope on each side of the dimeric capsid spike. We provide evidence that the HAstV capsid spike is a receptor-binding domain and that the antibody neutralizes HAstV by blocking virus attachment to cells. We identify patches of conserved amino acids that overlap the antibody epitope and may comprise a receptor-binding site. Our studies provide a foundation for the development of therapies to prevent and treat HAstV diarrheal disease.

    IMPORTANCEHuman astroviruses (HAstVs) infect nearly every person in the world during childhood and cause diarrhea, vomiting, and fever. Despite the prevalence of this virus, little is known about how antibodies in healthy adults protect them against reinfection. Here, we determined the crystal structure of a complex of the HAstV capsid protein and a virus-neutralizing antibody. We show that the antibody binds to the outermost spike domain of the capsid, and we provide evidence that the antibody blocks virus attachment to human cells. Importantly, our findings suggest that a subunit-based vaccine focusing the immune system on the HAstV capsid spike domain could be effective in protecting children against HAstV disease.

  11. Anti-idiotypic antibodies to poliovirus antibodies in commercial immunoglubulin preparations, human serum and milk.

    NARCIS (Netherlands)

    M. Hahn-Zoric; B. Carlsson; S. Jeansson; H.P. Ekre; A.D.M.E. Osterhaus (Albert); D. Roberton; L.A. Hanson

    1993-01-01

    textabstractOur previous studies have suggested that fetal antibody production can be induced by maternal antiidiotypic antibodies transferred to the fetus via the placenta. We tested commercial Ig, sera, and milk for the presence of anti-idiotypic antibodies to poliovirus type 1, using affinity

  12. Evading pre-existing anti-hinge antibody binding by hinge engineering

    Science.gov (United States)

    Kim, Hok Seon; Kim, Ingrid; Zheng, Linda; Vernes, Jean-Michel; Meng, Y. Gloria; Spiess, Christoph

    2016-01-01

    ABSTRACT Antigen-binding fragments (Fab) and F(ab′)2 antibodies serve as alternative formats to full-length anti-bodies in therapeutic and immune assays. They provide the advantage of small size, short serum half-life, and lack of effector function. Several proteases associated with invasive diseases are known to cleave antibodies in the hinge-region, and this results in anti-hinge antibodies (AHA) toward the neoepitopes. The AHA can act as surrogate Fc and reintroduce the properties of the Fc that are otherwise lacking in antibody fragments. While this response is desired during the natural process of fighting disease, it is commonly unwanted for therapeutic antibody fragments. In our study, we identify a truncation in the lower hinge region of the antibody that maintains efficient proteolytic cleavage by IdeS protease. The resulting neoepitope at the F(ab′)2 C-terminus does not have detectable binding of pre-existing AHA, providing a practical route to produce F(ab′)2 in vitro by proteolytic digestion when the binding of pre-existing AHA is undesired. We extend our studies to the upper hinge region of the antibody and provide a detailed analysis of the contribution of C-terminal residues of the upper hinge of human IgG1, IgG2 and IgG4 to pre-existing AHA reactivity in human serum. While no pre-existing antibodies are observed toward the Fab of IgG2 and IgG4 isotype, a significant response is observed toward most residues of the upper hinge of human IgG1. We identify a T225L variant and the natural C-terminal D221 as solutions with minimal serum reactivity. Our work now enables the production of Fab and F(ab′)2 for therapeutic and diagnostic immune assays that have minimal reactivity toward pre-existing AHA. PMID:27606571

  13. Facts and Fallacies of Kidd Antibodies: Experience in a Tertiary Care Hospital in North India.

    Science.gov (United States)

    Makroo, R N; Nayak, Sweta; Chowdhry, Mohit; Karna, Prashant

    2017-06-01

    We have analyzed the method used in our laboratory to detect the most elusive, clinically significant alloantibody: the Kidd alloantibodies and find the most convenient procedure. A retrospective analysis of the method used in our laboratory for determining Kidd alloantibodies from January 2013 to May 2015 was conducted. The details of the event that sensitized the patient for red cell antibody formation and procedure used to detect the alloantibody were retrieved from the departmental records. Of 405 red cell antibody identification cases, 24 (5.9 %) had Kidd antibody (anti-Jka in 12: 50 % cases; anti-Jkb in 4: 16.7 % cases; multiple antibodies in 8: 32 % cases). Thirteen of 24 patients (54.2 %) had autocontrol positive of which 6 cases needed adsorption procedures whereas antibody/ies could be identified without adsorption procedure in the remaining 7 cases. All the 7 cases had autocontrol of 1+ strength. Of the 11 patients (45.8 %) with autocontrol negative, the antibody was identified using solid phase in 7 cases whereas tube panels were also used in the remaining 4 cases. Kidd alloantibodies though deceptive can be identified by sensitive techniques like the solid phase and simple but laborious techniques using the tube cell panels. Depending upon the reaction strength of the autocontrol, the routine autoadsorption process may be skipped and tube cell enzyme treated cells or solid phase techniques be used to get the results.

  14. Effects of dexmedetomidine on H-FABP, CK-MB, cTnI levels, neurological function and near-term prognosis in patients undergoing heart valve replacement.

    Science.gov (United States)

    Wang, Zhi; Chen, Qiang; Guo, Hao; Li, Zhishan; Zhang, Jinfeng; Lv, Lei; Guo, Yongqing

    2017-12-01

    This study investigated the effects of dexmedetomidine on heart-type fatty acid binding protein (H-FABP), creatine kinase isoenzymes (CK-MB), and troponin I (cTnI) levels, neurological function and near-term prognosis in patients undergoing heart valve replacement. Patients undergoing heart valve replacement were randomly allocated to remifentanil anesthesia (control group, n=48) or dexmedetomidine anesthesia (observation group, n=48). Hemodynamic parameters were measured before anesthesia induction (T1), 1 min after intubation (T2), 10 min after start of surgery (T3), and on completion of surgery (T4). Levels of plasma H-FABP, CK-MB and cTnI were measured 10 min before anesthesia induction (C1), 10 min after start of surgery (C2), on completion of surgery (C3), 6 h after surgery (C4), and 24 h after surgery (C5). S100β protein and serum neuron-specific enolase (NSE) were detected 10 min before anesthesia induction (C1), and 24 h after surgery (C5). Neurological and cardiac function was evaluated 24 h after surgery. Incidence of cardiovascular adverse events was recorded for 1 year of follow-up. There were no significant differences in the average heart rate between the two groups during the perioperative period. The mean arterial pressure in the observation group was significantly lower than control group (PH-FABP, CK-MB and cTnI at C2, C3, C4 and C5, were significantly higher than C1, but significantly lower in the observation versus control group (P<0.05). Twenty-four hours after surgery, levels of S100β and NSE in both groups were higher than those before induction (P<0.05), but significantly lower in the observation versus control group (P<0.05). Twenty-four hours after surgery, neurological function scores were better, and myocardial contractility and arrhythmia scores significantly lower in the observation versus control group (P<0.05 for all). After follow-up for 1 year, incidence of cardiovascular adverse events was significantly lower in the observation

  15. Thoughts on identifiers

    CERN Document Server

    CERN. Geneva

    2005-01-01

    As business processes and information transactions have become an inextricably intertwined with the Web, the importance of assignment, registration, discovery, and maintenance of identifiers has increased. In spite of this, integrated frameworks for managing identifiers have been slow to emerge. Instead, identification systems arise (quite naturally) from immediate business needs without consideration for how they fit into larger information architectures. In addition, many legacy identifier systems further complicate the landscape, making it difficult for content managers to select and deploy identifier systems that meet both the business case and long term information management objectives. This presentation will outline a model for evaluating identifier applications and the functional requirements of the systems necessary to support them. The model is based on a layered analysis of the characteristics of identifier systems, including: * Functional characteristics * Technology * Policy * Business * Social T...

  16. Identifiability in stochastic models

    CERN Document Server

    1992-01-01

    The problem of identifiability is basic to all statistical methods and data analysis, occurring in such diverse areas as Reliability Theory, Survival Analysis, and Econometrics, where stochastic modeling is widely used. Mathematics dealing with identifiability per se is closely related to the so-called branch of ""characterization problems"" in Probability Theory. This book brings together relevant material on identifiability as it occurs in these diverse fields.

  17. High-Throughput Tools for Characterization of Antibody Epitopes

    DEFF Research Database (Denmark)

    Christiansen, Anders

    binders. In this study, phage display screenings were used to identify peptides that could inhibit a major toxin in cobra snake venom, α-cobratoxin. Peptide inhibitors were successfully identified. Importantly, HTS enabled the identification of toxin inhibitors that were not discovered by traditional...... phage display. Phage display coupled with HTS was again used in Chapter 3 in an attempt to map the epitopes of a therapeutic target injected into animals. The animals were immunized with a therapeutic target and the expectation was that they develop antibodies, which can be used in therapy. While...

  18. Monoclonal antibodies in pediatric allergy

    Directory of Open Access Journals (Sweden)

    Amelia Licari

    2015-10-01

    Full Text Available Production of monoclonal antibodies (mAbs involving human-mouse hybrid cells was first described in 1970s, but these biologics are now used for a variety of diseases including cancers, autoimmune disorders and allergic diseases. The aim of this article is to review current and future applications of mAbs, in particular focusing on anti-IgE therapy, in the field of pediatric allergy. Proceedings of the 11th International Workshop on Neonatology and Satellite Meetings · Cagliari (Italy · October 26th-31st, 2015 · From the womb to the adultGuest Editors: Vassilios Fanos (Cagliari, Italy, Michele Mussap (Genoa, Italy, Antonio Del Vecchio (Bari, Italy, Bo Sun (Shanghai, China, Dorret I. Boomsma (Amsterdam, the Netherlands, Gavino Faa (Cagliari, Italy, Antonio Giordano (Philadelphia, USA

  19. Update on antiphospholipid antibody syndrome

    Directory of Open Access Journals (Sweden)

    Michelle Remião Ugolini Lopes

    Full Text Available Summary Antiphospholipid syndrome (APS is an autoimmune disease characterized by antiphospholipid antibodies (aPL associated with thrombosis and/or pregnancy morbidity. Most APS events are directly related to thrombotic events, which may affect small, medium or large vessels. Other clinical features like thrombocytopenia, nephropathy, cardiac valve disease, cognitive dysfunction and skin ulcers (called non-criteria manifestations add significant morbidity to this syndrome and represent clinical situations that are challenging. APS was initially described in patients with systemic lupus erythematosus (SLE but it can occur in patients without any other autoimmune disease. Despite the autoimmune nature of this syndrome, APS treatment is still based on anticoagulation and antiplatelet therapy.

  20. [Radiolabeled antibodies for cancer treatment].

    Science.gov (United States)

    Barbet, Jacques; Chatal, Jean-François; Kraeber-Bodéré, Françoise

    2009-12-01

    The first treatment ever by radio-immunotherapy (RIT) was performed by William H. Beierwaltes in 1951 and was a success. Fifty years later, the main question is to find ways of extending the success of radiolabelled anti-CD20 antibodies in indolent non-Hodgkin's lymphoma to other forms of cancer. Solid tumours are much more radioresistant than lymphomas, but they respond to RIT if the lesions are small. Clinical situations of residual or minimal disease are thus the most likely to benefit from RIT in the adjuvant or consolidation settings. For disseminated disease, like leukemias or myelomas, the problem is different: beta- particles emitted by the radioactive atoms classically used for cancer treatment (iodine-131 or yttrium-90) disperse their energy in large volumes (ranges 1 mm to 1 cm) and are not very effective against isolated cells. Advances in RIT progress in two directions. One is the development of pretargeting strategies in which the antibody is not labelled but used to provide binding sites to small molecular weight radioactivity vectors (biotin, haptens). These techniques have been shown to increase tumour to non-target uptake ratios and anti-tumour efficacy has been demonstrated in the clinic. The other approach is the use of radionuclides adapted to the various clinical situations. Lutetium-177 or copper-67, because of the lower energy of their emission, their relatively long half-life and good gamma emission, may significantly improve RIT efficacy and acceptability. Beyond that, radionuclides emitting particles such as alpha particles or Auger electrons, much more efficient to kill isolated tumour cells, are being tested for RIT in the clinic. Finally, RIT should be integrated with other cancer treatment approaches in multimodality protocols. Thus RIT, now a mature technology, should enter a phase of well designed and focused clinical developments that may be expected to afford significant therapeutic advances.

  1. Applications of recombinant antibodies in plant pathology.

    Science.gov (United States)

    Ziegler, Angelika; Torrance, Lesley

    2002-09-01

    Summary Advances in molecular biology have made it possible to produce antibody fragments comprising the binding domains of antibody molecules in diverse heterologous systems, such as Escherichia coli, insect cells, or plants. Antibody fragments specific for a wide range of antigens, including plant pathogens, have been obtained by cloning V-genes from lymphoid tissue, or by selection from large naive phage display libraries, thus avoiding the need for immunization. The antibody fragments have been expressed as fusion proteins to create different functional molecules, and fully recombinant assays have been devised to detect plant viruses. The defined binding properties and unlimited cheap supply of antibody fusion proteins make them useful components of standardized immunoassays. The expression of antibody fragments in plants was shown to confer resistance to several plant pathogens. However, the antibodies usually only slowed the progress of infection and durable 'plantibody' resistance has yet to be demonstrated. In future, it is anticipated that antibody fragments from large libraries will be essential tools in high-throughput approaches to post-genomics research, such as the assignment of gene function, characterization of spatio-temporal patterns of protein expression, and elucidation of protein-protein interactions.

  2. Quantitative Changes In Antibodies Against Onchocercal Native ...

    African Journals Online (AJOL)

    Quantitative Changes In Antibodies Against Onchocercal Native Antigens Two Months Postivermectin Treatment Of Onchocerciasis Patients. ... Those without onchocercal skin disease, OSD (n=18) had a significant increase of 20.5±29.6%, with pre- and posttreatment values of 0.59±0.15 versus 0.68±0.13 for IgG antibody ...

  3. Anti-influenza M2e antibody

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M.

    2013-04-16

    Humanized recombinant and monoclonal antibodies specific for the ectodomain of the influenza virus M2 ion channel protein are disclosed. The antibodies of the invention have anti-viral activity and may be useful as anti-viral therapeutics and/or prophylactic/vaccine agents for inhibiting influenza virus replication and for treating individuals infected with influenza.

  4. Anti-influenza M2e antibody

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M [Santa Fe, NM

    2011-12-20

    Humanized recombinant and monoclonal antibodies specific for the ectodomain of the influenza virus M2 ion channel protein are disclosed. The antibodies of the invention have anti-viral activity and may be useful as anti-viral therapeutics and/or prophylactic/vaccine agents for inhibiting influenza virus replication and for treating individuals infected with influenza.

  5. Monoclonal Antibody Therapy for Advanced Neuroblastoma

    Science.gov (United States)

    NCI is sponsoring two clinical trials of a monoclonal antibody called ch14.18, in combination with other drugs, to see if the antibody may be helpful for children or young adults (up to age 21) with relapsed or refractory neuroblastoma.

  6. Serum Antiphospholipid Antibodies Among Healthy Adults In ...

    African Journals Online (AJOL)

    Background: Antiphospholipid antibodies have been associated with variety of conditions. There is no standard health associated reference values required for the interpretation of antiphospholipid antibodies result available among adults in North- eastern Nigeria and Nigeria in general. The aim of this study is to determine ...

  7. Radiolabeled antibodies for cancer imaging and therapy.

    Science.gov (United States)

    Barbet, Jacques; Bardiès, Manuel; Bourgeois, Mickael; Chatal, Jean-François; Chérel, Michel; Davodeau, François; Faivre-Chauvet, Alain; Gestin, Jean-François; Kraeber-Bodéré, Françoise

    2012-01-01

    Radiolabeled antibodies were studied first for tumor detection by single-photon imaging, but FDG PET stopped these developments. In the meantime, radiolabeled antibodies were shown to be effective in the treatment of lymphoma. Radiolabeling techniques are well established and radiolabeled antibodies are a clinical and commercial reality that deserves further studies to advance their application in earlier phase of the diseases and to test combination and adjuvant therapies including radiolabeled antibodies in hematological diseases. In solid tumors, more resistant to radiations and less accessible to large molecules such as antibodies, clinical efficacy remains limited. However, radiolabeled antibodies used in minimal or small-size metastatic disease have shown promising clinical efficacy. In the adjuvant setting, ongoing clinical trials show impressive increase in survival in otherwise unmanageable tumors. New technologies are being developed over the years: recombinant antibodies and pretargeting approaches have shown potential in increasing the therapeutic index of radiolabeled antibodies. In several cases, clinical trials have confirmed preclinical studies. Finally, new radionuclides, such as lutetium-177, with better physical properties will further improve the safety of radioimmunotherapy. Alpha particle and Auger electron emitters offer the theoretical possibility to kill isolated tumor cells and microscopic clusters of tumor cells, opening the perspective of killing the last tumor cell, which is the ultimate challenge in cancer therapy. Preliminary preclinical and preliminary clinical results confirm the feasibility of this approach.

  8. Determination of antiphospholipid antibodies and Thrombophilia in ...

    African Journals Online (AJOL)

    Background: Recurrent miscarriage is a critical problem in which many factors play a crucial role such as antiphospholipid antibodies (APA) and anticardiolipin antibodies (ACA). Recent studies pointed to a potential role of thrombophilias as a possible cause of recurrent miscarriage (RM). Objectives: This study was ...

  9. A novel polyclonal antibody against human cytomegalovirus ...

    African Journals Online (AJOL)

    Future research should be directed to epitope screening of synthetic HMCV peptides, which could help to understand HCMV infection and virus-neutralising antibodies more fully and to prepare HCMV vaccines and antiviral drugs. Key words: Human cytomegalovirus, AD169 strain, Towne strains, polyclonal antibody.

  10. Monoclonal antibodies reactive with hairy cell leukemia

    NARCIS (Netherlands)

    Visser, L; Shaw, A; Slupsky, J; Vos, H; Poppema, S

    Monoclonal antibodies reactive with hairy cell leukemia were developed to aid in the diagnosis of this subtype of B cell chronic lymphocytic leukemia and to gain better insight into the origin of hairy cells. Three antibodies were found to be of value in the diagnosis of hairy cell leukemia.

  11. [Irregular antibody testing during pregnancy in Tunisia: clinical study of 5369 women].

    Science.gov (United States)

    Rekik, T; Ben Amor, I; Louati, N; Rekik, H; Menif, H; Gargouri, J

    2012-04-01

    To evaluate the prevalence of alloimmunization in women followed in an obstetrical environment in Tunisia, to identify the specificities of antibodies found and to determine factors that could influence the appearance of this immunization. We proceeded to a retrospective analysis of search for irregular antibodies in women followed up in obstetrical environment over nine consecutive years (2000-2008). The panel was officially defined and produced by the Regional Centre for Blood Transfusion in Sfax (Tunisia). Overall 5369 women benefited from 6575 antibody testing (average: 1.22; extremes: 1-14). The results were positive for 278 women (5.17 %), allowing to identify 216 antibodies or associations of antibodies. Among identified antibodies, those immune were found in 198 women. The rate of alloimmunization was 3.68 % (198/5369). The majority of the antibodies found was anti-Rh1, isolated or associated with another antibody, in 84.3 % of the total immunized women. The immunization of women according to the number of gestations showed a significant increasing rate ranging from 2.34 % for a first gestation to 5.27 % for four gestations or more. In addition, a significant difference was also noted between the rate of immunization in women who had received anti-Rh1 immunoglobulin and those who had not. Anti-Rh1 immunization is the most frequent in the population of studied women. This could denote of an insufficiency in pregnancies follow-up and immunoprophylaxis protocols. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  12. Many Routes to an Antibody Heavy-Chain CDR3: Necessary, Yet Insufficient, for Specific Binding

    Science.gov (United States)

    D’Angelo, Sara; Ferrara, Fortunato; Naranjo, Leslie; Erasmus, M. Frank; Hraber, Peter; Bradbury, Andrew R. M.

    2018-01-01

    Because of its great potential for diversity, the immunoglobulin heavy-chain complementarity-determining region 3 (HCDR3) is taken as an antibody molecule’s most important component in conferring binding activity and specificity. For this reason, HCDR3s have been used as unique identifiers to investigate adaptive immune responses in vivo and to characterize in vitro selection outputs where display systems were employed. Here, we show that many different HCDR3s can be identified within a target-specific antibody population after in vitro selection. For each identified HCDR3, a number of different antibodies bearing differences elsewhere can be found. In such selected populations, all antibodies with the same HCDR3 recognize the target, albeit at different affinities. In contrast, within unselected populations, the majority of antibodies with the same HCDR3 sequence do not bind the target. In one HCDR3 examined in depth, all target-specific antibodies were derived from the same VDJ rearrangement, while non-binding antibodies with the same HCDR3 were derived from many different V and D gene rearrangements. Careful examination of previously published in vivo datasets reveals that HCDR3s shared between, and within, different individuals can also originate from rearrangements of different V and D genes, with up to 26 different rearrangements yielding the same identical HCDR3 sequence. On the basis of these observations, we conclude that the same HCDR3 can be generated by many different rearrangements, but that specific target binding is an outcome of unique rearrangements and VL pairing: the HCDR3 is necessary, albeit insufficient, for specific antibody binding. PMID:29568296

  13. Many Routes to an Antibody Heavy-Chain CDR3: Necessary, Yet Insufficient, for Specific Binding.

    Science.gov (United States)

    D'Angelo, Sara; Ferrara, Fortunato; Naranjo, Leslie; Erasmus, M Frank; Hraber, Peter; Bradbury, Andrew R M

    2018-01-01

    Because of its great potential for diversity, the immunoglobulin heavy-chain complementarity-determining region 3 (HCDR3) is taken as an antibody molecule's most important component in conferring binding activity and specificity. For this reason, HCDR3s have been used as unique identifiers to investigate adaptive immune responses in vivo and to characterize in vitro selection outputs where display systems were employed. Here, we show that many different HCDR3s can be identified within a target-specific antibody population after in vitro selection. For each identified HCDR3, a number of different antibodies bearing differences elsewhere can be found. In such selected populations, all antibodies with the same HCDR3 recognize the target, albeit at different affinities. In contrast, within unselected populations, the majority of antibodies with the same HCDR3 sequence do not bind the target. In one HCDR3 examined in depth, all target-specific antibodies were derived from the same VDJ rearrangement, while non-binding antibodies with the same HCDR3 were derived from many different V and D gene rearrangements. Careful examination of previously published in vivo datasets reveals that HCDR3s shared between, and within, different individuals can also originate from rearrangements of different V and D genes, with up to 26 different rearrangements yielding the same identical HCDR3 sequence. On the basis of these observations, we conclude that the same HCDR3 can be generated by many different rearrangements, but that specific target binding is an outcome of unique rearrangements and VL pairing: the HCDR3 is necessary, albeit insufficient, for specific antibody binding.

  14. Production and characterization of peptide antibodies

    DEFF Research Database (Denmark)

    Trier, Nicole Hartwig; Hansen, Paul Robert; Houen, Gunnar

    2012-01-01

    Proteins are effective immunogens for generation of antibodies. However, occasionally the native protein is known but not available for antibody production. In such cases synthetic peptides derived from the native protein are good alternatives for antibody production. These peptide antibodies...... are powerful tools in experimental biology and are easily produced to any peptide of choice. A widely used approach for production of peptide antibodies is to immunize animals with a synthetic peptide coupled to a carrier protein. Very important is the selection of the synthetic peptide, where factors...... such as structure, accessibility and amino acid composition are crucial. Since small peptides tend not to be immunogenic, it may be necessary to conjugate them to carrier proteins in order to enhance immune presentation. Several strategies for conjugation of peptide-carriers applied for immunization exist...

  15. Antiphospholipid antibody: laboratory, pathogenesis and clinical manifestations

    Directory of Open Access Journals (Sweden)

    T. Ziglioli

    2011-06-01

    Full Text Available Antiphospholipid antibodies (aPL represent a heterogeneous group of antibodies that recognize various antigenic targets including beta2 glycoprotein I (β2GPI, prothrombin (PT, activated protein C, tissue plasminogen activator, plasmin and annexin A2. The most commonly used tests to detect aPL are: lupus anticoagulant (LAC, a functional coagulation assay, anticardiolipin antibody (aCL and anti-β2GPI antibody (anti-β2GPI, which are enzyme-linked immunoassay (ELISA. Clinically aPL are associated with thrombosis and/or with pregnancy morbidity. Apparently aPL alone are unable to induce thrombotic manifestations, but they increase the risk of vascular events that can occur in the presence of another thrombophilic condition; on the other hand obstetrical manifestations were shown to be associated not only to thrombosis but mainly to a direct antibody effect on the trophoblast.

  16. Antibodies against chromosomal beta-lactamase

    DEFF Research Database (Denmark)

    Giwercman, B; Rasmussen, J W; Ciofu, Oana

    1994-01-01

    A murine monoclonal anti-chromosomal beta-lactamase antibody was developed and an immunoblotting technique was used to study the presence of serum and sputum antibodies against Pseudomonas aeruginosa chromosomal group 1 beta-lactamase in patients with cystic fibrosis (CF). The serum antibody...... response was studied with serum samples collected in 1992 from 56 CF patients in a cross-sectional study and with serum samples from 18 CF patients in a longitudinal study. Anti-beta-lactamase immunoglobulin G antibodies were present in all of the serum samples from the patients with chronic...... bronchopulmonary P. aeruginosa infection (CF + P) but in none of the CF patients with no or intermittent P. aeruginosa infection. Anti-beta-lactamase antibodies were present in serum from CF + P patients after six antipseudomonal courses (median) and correlated with infection with a beta-lactam-resistant strain...

  17. Onconeural antibodies: improved detection and clinical correlations.

    Science.gov (United States)

    Storstein, Anette; Monstad, Sissel Evy; Haugen, Mette; Mazengia, Kibret; Veltman, Dana; Lohndal, Emilia; Aarseth, Jan; Vedeler, Christian

    2011-03-01

    Onconeural antibodies are found in many patients with paraneoplastic neurological syndromes (PNS) and define the disease as paraneoplastic. The study describes the presence of onconeural antibodies and PNS in 555 patients with neurological symptoms and confirmed cancer within five years, and compares the diagnostic accuracy of different antibody assays (immunoprecipitation, immunofluorescence and immunoblot). Onconeural antibodies were found in 11.9% of the patients by immunoprecipitation, in 7.0% by immunofluorescence and in 6.3% by immunoblot. PNS were present in 81.8% of the cancer patients that were seropositive by immunoprecipitation. Immunofluorescence and immunoblot failed to detect onconeural antibodies in almost one third of the PNS cases. Copyright © 2010 Elsevier B.V. All rights reserved.

  18. [Advance of clinical study on immune thrombocytopenia caused by irregular antibodies].

    Science.gov (United States)

    Zhu, Lin-Lin; Zhang, Chen-Guang

    2011-06-01

    The platelet antibodies mainly include platelet-specific and related antibodies, which belong to irregular antibodies. They are produced by autoimmune, drug-induced or isoimmunization (such as pregnancy, blood transfusion and so on), the irregular IgG and/or IgM antibodies produce and lead to platelet transfusion refractoriness (PTR), post-transfusion purpura (PTP) and isoimmune neonatal thrombocytopenic purpura (INTP), idiopathic thrombocytopenic purpura and so on. It is very necessary to screen and identify the irregular antibodies before blood transfusion or parturition. Except some serological detections should be done first, flow cytometry and molecular biological techniques such as PCR and PCR-SSP are applied to detect the difficult-matching patients' genotypes and fetal genotypes in order to further predict fetal INTP and to provide the right blood for difficult-matching patients, therefore, some measures must be taken early for prevention and treatment of immune thrombocytopenic purpura. In this review, the production, typing and laboratory tests of irregular antibodies, as well as the pathogenesis and clinical symptoms of diseases caused by irregular antibodies, and the current progress are summarized.

  19. Soluble HIV-1 envelope immunogens derived from an elite neutralizer elicit cross-reactive V1V2 antibodies and low potency neutralizing antibodies.

    Directory of Open Access Journals (Sweden)

    Sara Carbonetti

    Full Text Available We evaluated four gp140 Envelope protein vaccine immunogens that were derived from an elite neutralizer, subject VC10042, whose plasma was able to potently neutralize a wide array of genetically distinct HIV-1 isolates. We sought to determine whether soluble Envelope proteins derived from the viruses circulating in VC10042 could be used as immunogens to elicit similar neutralizing antibody responses by vaccination. Each gp140 was tested in its trimeric and monomeric forms, and we evaluated two gp140 trimer vaccine regimens in which adjuvant was supplied at all four immunizations or at only the first two immunizations. Interestingly, all four Envelope immunogens elicited high titers of cross-reactive antibodies that recognize the variable regions V1V2 and are potentially similar to antibodies linked with a reduced risk of HIV-1 acquisition in the RV144 vaccine trial. Two of the four immunogens elicited neutralizing antibody responses that neutralized a wide array of HIV-1 isolates from across genetic clades, but those responses were of very low potency. There were no significant differences in the responses elicited by trimers or monomers, nor was there a significant difference between the two adjuvant regimens. Our study identified two promising Envelope immunogens that elicited anti-V1V2 antibodies and broad, but low potency, neutralizing antibody responses.

  20. Children with multiphasic disseminated encephalomyelitis and antibodies to the myelin oligodendrocyte glycoprotein (MOG): Extending the spectrum of MOG antibody positive diseases.

    Science.gov (United States)

    Baumann, Matthias; Hennes, Eva-Maria; Schanda, Kathrin; Karenfort, Michael; Kornek, Barbara; Seidl, Rainer; Diepold, Katharina; Lauffer, Heinz; Marquardt, Iris; Strautmanis, Jurgis; Syrbe, Steffen; Vieker, Silvia; Höftberger, Romana; Reindl, Markus; Rostásy, Kevin

    2016-12-01

    Myelin oligodendrocyte glycoprotein (MOG) antibodies have been described in children with acute disseminated encephalomyelitis (ADEM), recurrent optic neuritis, neuromyelitis optica spectrum disorders and more recently in children with multiphasic disseminated encephalomyelitis (MDEM). To delineate the clinical, cerebrospinal fluid (CSF) and radiological features of paediatric MDEM with MOG antibodies. Clinical course, serum antibodies, CSF, magnetic resonance imaging (MRI) studies and outcome of paediatric MDEM patients were reviewed. A total of 8 children with two or more episodes of ADEM were identified from a cohort of 295 children with acute demyelinating events. All children had persisting MOG antibodies (median titre: 1:1280). All ADEM episodes included encephalopathy, polyfocal neurological signs and a typical MRI. Apart from ADEM episodes, three children had further clinical attacks without encephalopathy. Median age at initial presentation was 3 years (range: 1-7 years) and median follow-up 4 years (range: 1-8 years). New ADEM episodes were associated with new neurological signs and new MRI lesions. Clinical outcome did range from normal (four of the eight) to mild or moderate impairment (four of the eight). A total of four children received monthly immunoglobulin treatment during the disease course. Children with MDEM and persisting MOG antibodies constitute a distinct entity of relapsing demyelinating events and extend the spectrum of MOG antibody-associated diseases. © The Author(s), 2016.