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Sample records for chain fv constructs

  1. Homology modelling and bivalent single-chain Fv construction of anti-HepG2 single-chain immunoglobulin Fv fragments from a phage display library

    Indian Academy of Sciences (India)

    Ming Ni; Bing Yu; Y U Huang; Zhenjie Tang; Ping Lei; Xin Shen; Wei Xin; Huifen Zhu; Guanxin Shen

    2008-12-01

    We prepared single-chain immunoglobulin Fv fragments (scFv) SLH10 specific for the HepG2 cell line after biopanning from a large human-naïve phage display library (Griffin. 1 Library). The three-dimensional (3D) structure of SLH10 was modelled by the Insight II molecule simulation software. The structure was refined using the molecular dynamics method. The structures with the least steric clashes and lowest energy were determined finally. The optimized structures of heavy (VH) and light (VL) variable chains of SLH10 scFv were obtained. Then SLH10 bivalent single-chain Fv (BsFv) was constructed that would be suitable for high-affinity targeting. SLH10 BsFv was generated by linking scFvs together and identified by sequencing. Its expression products were confirmed by western blot analysis. The relative molecular masses of scFv and BsFv were approximately 30 kDa and 60 kDa, respectively. Flow cytometry revealed that SLH10 BsFv bound the selected cell lines with greater signal intensity than the parental scFv. The improved antigen binding of SLH10 BsFv may be useful for immunodiagnostics or targeted gene therapy for liver cancer.

  2. Construction, expression and tumor targeting of a single-chain Fv against human colorectal carcinoma

    Institute of Scientific and Technical Information of China (English)

    Jin Fang; Hong-Bin Jin; Jin-Dan Song

    2003-01-01

    AIM: A single-chain antibody fragment, ND-1scFv, against human colorectal carcinoma was constructed and expressed in E.coli, and its biodistribution and pharmacokinetic properties were studied in mice bearing tumor.METHODS: VH and VL genes were amplified from hybridoma cell IC-2, secreting monoclonal antibody ND-1, by RT-PCR,and connected by linker (Gly4Ser)3 to form scFv gene, which was cloned into expression vector pET 28a(+) and finally expressed in E.coli. The expressed product ND-1scFv was purified by metal affinity chromatography using Ni-NTA, its purity and biological activity were determined using SDSPAGE and ELISA. ND-1scFv was labeled with 99mTc, and then injected into mice bearing colorectal carcinoma xenograft for phamacokinetic study in vivo.RESULTS: SDS-PAGE analysis showed that the relative molecular weight of recombinant protein was 30kDa with purity of 94%. ELIAS assay revealed that ND-1scFv retained the immunoactivity of parent mAb, being capable of binding specifically to human colorectal carcinoma cell line expressing associated antigen. Radiolabeled ND-1scFv exhibited rapid tumor targeting, with specific distribution in mice bearing colorectal carcinoma xenograft observed as early as 1 h following injection. In vivo pharmacokinetic studies also demonstrated that ND-1scFv had very rapid plasma clearance (T1/2α of 5.7 min, T1/2β of 2.6 h).CONCLUSION: ND-1scFv shows significant immunoactivity,and better pharmacokinetic and biodistribution characteristics compared with intact mAbs, demonstrating the possibility as a carrier for tumor-imaging.

  3. Single Chain Fv Constructs of Anti-Ganglioside GD2 Antibodies for Radioimaging and Radioimmunotherapy

    International Nuclear Information System (INIS)

    of its broad and usually homogeneous distribution in human solid tumors, and most importantly, their absence on cell membranes of normal human tissues. In separate experiments, we have shown that T-cells transduced with the herpes simplex viral thymidine kinase (HSV-tk) gene can be radiolabeled with 131I-FIAU to a safe nuclear radiation dose. Using a dicistronic construct we are inserting chimeric immune receptor plus HSV-tk into T-cells to allow such their trafficking to be radioactively monitored. We plan to study the role of cytokines, chemoreceptors and CD4 helper T-cells in recruiting CD8+ transduced T-cells to the tumor site. These studies should provide us with an adoptive cell therapy approach to target cytotoxicity to human tumors, and a lymphocyte tracking tool to study delivery to the tumor sites, to determine if they proliferate locally and/or recirculate. Such pharmacologic information is crucial for optimizing gene-modified T-cells in future clinical trials. Single chain FV constructs of anti-ganglioside GD2 antibodies for radioimaging

  4. Construction and structural modeling of a single-chain Fv-asparaginase fusion protein resistant to proteolysis.

    Science.gov (United States)

    Guo, L; Wang, J; Qian, S; Yan, X; Chen, R; Meng, G

    2000-11-20

    In this study, we construct a fusion protein composed of L-asparaginase (ASNase; from Escherichia coli AS 1.357) and a protective single-chain Fv (scFv), which was selected from a phage-display scFv library from our previous studies. The antibody moiety of the fusion protein was fused to the N-terminus of the enzyme moiety via a linker peptide, (Gly(4)Ser)(6). Recombinant plasmid pET-SLA was constructed to express scFv-ASNase fusion to high levels in E. coli and the expressed product was found to form inclusion bodies. We obtained a soluble fusion protein by refolding and purification. The soluble fusion protein exhibited about 82% of the enzymatic activity of the native ASNase at the same molar concentration, and had a K(m) value similar to that of the native enzyme for the substrate L-asparagine. Importantly, the fusion protein was more stable than native ASNase. In addition: (1) following treatment with trypsin, alpha-chymotrypsin, and rennet, at 37 degrees C for 30 min, scFv-ASNase fusion retained 94.0%, 88.8%, and 84.5% of its original activity, respectively, whereas native ASNase became inactive; and (2) ScFv-ASNase fusion had a much longer in vitro half-life (9 h) in serum than the native enzyme (2 h). The three-dimensional structure of the fusion protein was obtained by modeling with the Homology and Discover modules of the INSIGHT II software package. On the basis of the structural evidence and biochemical properties, we propose that the scFv moiety of the fusion protein may confer ASNase moiety resistance to proteolysis as a result of both steric hindrance and a change in the electrostatic surface of the enzyme. PMID:11005928

  5. Generation of AcGFP fusion with single-chain Fv selected from a phage display library constructed from mice hyperimmunized against 5-methyl 2'-deoxycytidine.

    Science.gov (United States)

    Ohshima, Motohiro; Inoue, Kazuyuki; Hayashi, Hideki; Tsuji, Daiki; Mizugaki, Michinao; Itoh, Kunihiko

    2010-11-01

    DNA methylation is involved in many diseases such as cancer and autoimmunity. We generated recombinant single-chain Fv (scFv) antibodies against 5-methyl-2'-deoxycytidine (m(5)dCyd) using phage display technology and a hyperimmunized mouse, and the scFv of most interest were constructed as fusion proteins with green fluorescent protein obtained from Aequorea coerulescens GFP (AcGFP). Using RNA isolated from mouse spleens, we constructed a scFv library consisting of λ light chains. The scFv library was selected against m(5)Cyd-BSA and enriched through four rounds of panning. The scFv library was concentrated about 390-fold and an individual clone was reacted with m(5)Cyd-BSA. Two scFvs with high reactivity for m(5)Cyd-BSA termed 1-2 and 1-12 were produced. Furthermore, methylated DNA-binding activities of the scFvs were confirmed using an indirect immunofluorescence assay. Additionally, N- and C-terminal scFv 1-2 fusion with AcGFP were constructed, and we observed the N-terminal AcGFP exhibited much higher fluorescence intensity than the C-terminal fusions. The AcGFP-scFv 1-2 modified N-terminus of scFv with AcGFP had high fluorescence intensity, but the scFv 1-2-AcGFP modified C-terminus of scFv with AcGFP had low fluorescence intensity. The cross-reactivity of AcGFP-scFv 1-2 was similar to scFv 1-2, and thus, AcGFP-scFv 1-2 could be used in a direct immunofluorescence assay. The scFv fusion proteins may be useful for the detection and quantification of cellular methylated DNA in various specimens. PMID:20876190

  6. Pharmacokinetics and biodistribution of 177Lu-labeled multivalent single-chain Fv construct of the pancarcinoma monoclonal antibody CC49

    International Nuclear Information System (INIS)

    Lutetium-177 (177Lu) is a radionuclide of interest for radioimmunoimaging (RII) and radioimmunotherapy (RIT) on account of its short half-life (161 h) and the ability to emit both β and γ radiation. Single-chain Fv (scFv) constructs have shown advancement in cancer diagnosis and therapy due to the pharmacokinetics advantage and seem to be intriguing tools in oncology. The objective of this study was to evaluate the pharmacokinetics and biodistribution characteristics of the 177Lu-labeled tetravalent scFv of CC49 MAb and intact CC49 IgG in vivo. Conjugation and labeling conditions of multivalent scFv with 177Lu were optimized without affecting integrity and immunoreactivity. For this purpose, multivalent scFv constructs dimer, sc(Fv)2; tetramer, [sc(Fv)2]2 of the MAb CC49 were expressed as secretory proteins in Pichia pastoris. The purified scFv constructs and IgG form of CC49 were conjugated with a bifunctional chelating agent, ITCB-DTPA, and labeled with 177Lu. The comparative biodistribution, blood clearance, and tumor-targeting characteristics of 177Lu-labeled tetravalent [sc(Fv)2 ]2 construct of CC49 MAb and intact CC49 IgG were investigated in the athymic mice bearing LS-174T xenografts. Approximately, 90% of 177Lu incorporation was achieved using ITCB-DTPA chelator, and the labeled immunoconjugates maintained integrity and immunoreactivity. Blood clearance studies demonstrated an alpha half-life (t1/2 α) of 177Lu-labeled [sc(Fv)2 ]2 and IgG of CC49 at 4.40 and 9.50 min and a beta half-life (t1/2 β) at 375 and 2,193 min, respectively. At 8 h post administration, the percent of the injected dose accumulated/gram (%ID/g) of the LS-174T tumor was 6.4 ±1.3 and 8.9 ±0.6 for 177Lu-labeled [sc(Fv)2 ]2 and IgG of CC49, respectively, in the absence of l-lysine. The corresponding values were 8.0 ±0.6 and 8.4 ±1.2 in the presence of l-lysine. Renal accumulation of [sc(Fv)2 ]2 was significantly (p <0.005) reduced in the presence of l-lysine. (orig.)

  7. Pharmacokinetics and biodistribution of {sup 177}Lu-labeled multivalent single-chain Fv construct of the pancarcinoma monoclonal antibody CC49

    Energy Technology Data Exchange (ETDEWEB)

    Chauhan, Subhash C.; Jain, Maneesh; Moore, Erik D.; Wittel, Uwe A.; Batra, Surinder K. [University of Nebraska Medical Center, Department of Biochemistry and Molecular Biology, Omaha, NE (United States); Li, Jing; Gwilt, Peter R. [University of Nebraska Medical Center, College of Pharmacy, Omaha, NE (United States); Colcher, David [Beckman Research Institute at City of Hope National Medical Center, Department of Radioimmunotherapy, Duarte, CA (United States)

    2005-03-01

    Lutetium-177 ({sup 177}Lu) is a radionuclide of interest for radioimmunoimaging (RII) and radioimmunotherapy (RIT) on account of its short half-life (161 h) and the ability to emit both {beta} and {gamma} radiation. Single-chain Fv (scFv) constructs have shown advancement in cancer diagnosis and therapy due to the pharmacokinetics advantage and seem to be intriguing tools in oncology. The objective of this study was to evaluate the pharmacokinetics and biodistribution characteristics of the {sup 177}Lu-labeled tetravalent scFv of CC49 MAb and intact CC49 IgG in vivo. Conjugation and labeling conditions of multivalent scFv with {sup 177}Lu were optimized without affecting integrity and immunoreactivity. For this purpose, multivalent scFv constructs dimer, sc(Fv){sub 2}; tetramer, [sc(Fv){sub 2}]{sub 2} of the MAb CC49 were expressed as secretory proteins in Pichia pastoris. The purified scFv constructs and IgG form of CC49 were conjugated with a bifunctional chelating agent, ITCB-DTPA, and labeled with {sup 177}Lu. The comparative biodistribution, blood clearance, and tumor-targeting characteristics of {sup 177}Lu-labeled tetravalent [sc(Fv){sub 2} ]{sub 2} construct of CC49 MAb and intact CC49 IgG were investigated in the athymic mice bearing LS-174T xenografts. Approximately, 90% of {sup 177}Lu incorporation was achieved using ITCB-DTPA chelator, and the labeled immunoconjugates maintained integrity and immunoreactivity. Blood clearance studies demonstrated an alpha half-life (t{sub 1/2} {alpha}) of {sup 177}Lu-labeled [sc(Fv){sub 2} ]{sub 2} and IgG of CC49 at 4.40 and 9.50 min and a beta half-life (t{sub 1/2} {beta}) at 375 and 2,193 min, respectively. At 8 h post administration, the percent of the injected dose accumulated/gram (%ID/g) of the LS-174T tumor was 6.4 {+-}1.3 and 8.9 {+-}0.6 for {sup 177}Lu-labeled [sc(Fv){sub 2} ]{sub 2} and IgG of CC49, respectively, in the absence of l-lysine. The corresponding values were 8.0 {+-}0.6 and 8.4 {+-}1.2 in the

  8. Baculovirus display of single chain antibody (scFv using a novel signal peptide

    Directory of Open Access Journals (Sweden)

    Gonzalez Gaëlle

    2010-11-01

    Full Text Available Abstract Background Cells permissive to virus can become refractory to viral replication upon intracellular expression of single chain fragment variable (scFv antibodies directed towards viral structural or regulatory proteins, or virus-coded enzymes. For example, an intrabody derived from MH-SVM33, a monoclonal antibody against a conserved C-terminal epitope of the HIV-1 matrix protein (MAp17, was found to exert an inhibitory effect on HIV-1 replication. Results Two versions of MH-SVM33-derived scFv were constructed in recombinant baculoviruses (BVs and expressed in BV-infected Sf9 cells, N-myristoylation-competent scFvG2/p17 and N-myristoylation-incompetent scFvE2/p17 protein, both carrying a C-terminal HA tag. ScFvG2/p17 expression resulted in an insoluble, membrane-associated protein, whereas scFvE2/p17 was recovered in both soluble and membrane-incorporated forms. When coexpressed with the HIV-1 Pr55Gag precursor, scFvG2/p17 and scFvE2/p17 did not show any detectable negative effect on virus-like particle (VLP assembly and egress, and both failed to be encapsidated in VLP. However, soluble scFvE2/p17 isolated from Sf9 cell lysates was capable of binding to its specific antigen, in the form of a synthetic p17 peptide or as Gag polyprotein-embedded epitope. Significant amounts of scFvE2/p17 were released in the extracellular medium of BV-infected cells in high-molecular weight, pelletable form. This particulate form corresponded to BV particles displaying scFvE2/p17 molecules, inserted into the BV envelope via the scFv N-terminal region. The BV-displayed scFvE2/p17 molecules were found to be immunologically functional, as they reacted with the C-terminal epitope of MAp17. Fusion of the N-terminal 18 amino acid residues from the scFvE2/p17 sequence (N18E2 to another scFv recognizing CD147 (scFv-M6-1B9 conferred the property of BV-display to the resulting chimeric scFv-N18E2/M6. Conclusion Expression of scFvE2/p17 in insect cells using a BV

  9. [Construction and panning of scFv phage display library against recombinant interleukin 4 receptor].

    Science.gov (United States)

    Yang, Guangyong; Guo, Haitao; Liu, Ximing; He, Guangzhi; Tian, Weiyi; Cai, Kun; Wang, Ping; Wang, Wenjia

    2016-06-01

    Objective To construct the recombinant human interleukin 4 receptor (rhIL-4R) single-chain Fv (scFv) antibody library by phage display technique to obtain the anti-IL-4R scFv clones selected from the library. Methods Total RNA was extracted from splenocytes of the BALB/c mice immunized with rhIL-4R. Complementary DNA fragments of variable heavy (VH) and variable light (VL) chains of the antibodies were prepared by reverse transcription PCR and assembled into scFv by splice overlap extension PCR (SOE-PCR). Both scFv and the pCANTAB5E vector were respectively double-digested with restriction endonuclease Sfi I and Not I, connected with T4 ligase, and then transformed into the competent cells E.coli TG1; it was cultured in medium to obtain the phage scFv antibody library; after three rounds of enrichment and panning, the specific antigen scFv with high affinity was selected for the sequencing. Results After three rounds of panning, we obtained a diversity of approximately 2×10(8) anti-rhIL-4R scFv antibody library. Sequencing analysis of one positive clone showed that the anti-rhIL-4R scFv was 741 bp and coded 247 amino acids. The analysis of VBASE2 database indicated that VH and VL gene sequences of anti-rhIL-4R protein all had three complementarity determining regions and four backbone areas.Conclusion The anti-rhIL-4R scFv was obtained from the scFv antibody library. PMID:27371853

  10. Construction of Human ScFv Phage Display Library against Ovarian Tumor

    Institute of Scientific and Technical Information of China (English)

    XIA Jinsong; BI Hao; YAO Qin; QU Shen; ZONG Yiqiang

    2006-01-01

    In order to construct a single chain fragment variable (ScFv) phage display library against ovarian tumor, by using RT-PCR, the human heavy chain variable region genes (VH) and light chain variable region genes (VL) were amplified from lymphocytes of ovarian tumor patients and subsequently assembled into ScFv genes by SOE. The resulting ScFv genes were electrotransformed into E.coli TG1 and amplified with the co-infection of helper phage M13KO7 to obtain phage display library. The capacity and titer of the resulting library were detected. The phage antibody library with a capacity of approximately 3 × 109 cfu/μg was obtained. After amplification with helper phage, the titer of antibody library reached 5 × 1012 cfu/mL. Human ScFv library against ovarian tumor was constructed successfully, which laid a foundation for the screening of ovarian tumor specific ScFv for the radioimmunoimaging diagnosis of ovarian tumor.

  11. Negative effects of a disulfide bond mismatch in anti-rabies G protein single-chain antibody variable fragment FV57.

    Science.gov (United States)

    Duan, Ye; Gu, Tiejun; Zhang, Xizhen; Jiang, Chunlai; Yuan, Ruosen; Li, Zhuang; Wang, Dandan; Chen, Xiaoxu; Wu, Chunlai; Chen, Yan; Wu, Yongge; Kong, Wei

    2014-06-01

    Rabies virus (RV) causes a fatal infectious disease requiring efficient post-exposure prophylaxis (PEP), which includes a rabies vaccine and rabies immunoglobulin (RIG). The single-chain antibody variable fragment (scFv), a small engineered antibody fragment derived from an antibody variable heavy chain and light chain, has the potential to replace the current application of RIG. In previous studies, we constructed and evaluated an anti-rabies virus G protein scFv (FV57) based on the monoclonal antibody CR57. Of the five cysteines in FV57, four are linked in intra-chain disulfide bonds (Cys-VH28/Cys-VH98 and Cys-VL16/Cys-VL84), and one is free (Cys-VL85). However, the thiol in Cys-VL85 neighboring Cys-VL84 in the CDR3 of the light chain is likely to mismatch with the thiol in Cys-VL16 during the renaturing process. In order to study effects of the mismatched disulfide bond, Cys-VL85 and Cys-VL84 of FV57 were mutated to serine to construct mutants FV57(VL85S) and FV57(VL84S). Furthermore, the disulfide bonds in the light chain of FV57, FV57(VL85S) and FV57(VL84S) were deleted by mutating Cys-VL16 to serine. All mutants were prepared and evaluated along with the original FV57. The results indicated that the mismatched disulfide bond of FV57 linking the light chain FR1 and CDR3 would confer deleterious negative effects on its activity against RV, likely due to spatial hindrance in the light chain CDR3. Moreover, avoidance of the disulfide bond mismatch provided an additional 30% protective efficacy against RV infection in the mouse RV challenge model. Thus, modifications of FV57 to eliminate the disulfide bond mismatch may provide a candidate therapeutic agent for effective PEP against rabies. PMID:24598312

  12. Formation of disulfide bridges by a single-chain Fv antibody in the reducing ectopic environment of the plant cytosol

    NARCIS (Netherlands)

    Schouten, A.; Roosien, J.; Bakker, J.; Schots, A.

    2002-01-01

    Disulfide bridge formation in the reducing environment of the cytosol is considered a rare event and is mostly linked to inactivation of protein activity. In this report the in vivo redox state of a single-chain Fv (scFv) antibody fragment in the plant cytosol was investigated. The scFv antibody fra

  13. High efficient mammalian expression and secretion of a functional humanized single-chain Fv/human interleukin-2 molecules

    Institute of Scientific and Technical Information of China (English)

    Yue-Chun Shen; Xue-Hao Wang; Xiao-Ming Wang; Zao-Lai Chen; Xi-Ping Shen; Chao-Chen Zhao; Jun Li

    2006-01-01

    AIM: To construct and produce a recombinant bispecific humanized single-chain Fv (sFv) /Interleukin-2 (IL-2)fusion protein by using mammalian cells.METHODS: The sFv/IL-2 protein was genetically engineered, and transfected to mammalian cells to determine whether the mammalian protein folding machinery can produce and secrete active sFv/IL-2 with high efficiency.RESULTS: The fusion protein was constructed and high efficiently expressed with yields up to 102 ± 4.2 mg/L in culture supernatant of the stably transfected 293cell line. This recombinant fusion protein consisted of humanized variable heavy (VH) and light (VL) domains of monoclonal antibody (mAb) 520C9 directed against the human HER-2/neu (c-erbB2) proto-oncogene product p185, and human IL-2 connected by polypeptide linker. The fusion protein was shown to retain the immunostimulatory activities of IL-2 as measured by IL-2-dependent cell proliferation and cytotoxicity assays.In addition to its IL-2 activities, this fusion protein also possessed antigen-binding specificity against p185, as determined by indirect ELISA using p185 positive SKOV 3ip1 cells.CONCLUSION: The large-scale preparation of the recombinant humanized sFv antibody/IL-2 fusion protein is performed with 293 cells. The recombinant humanized sFv antibody/IL-2 fusion protein may provide an effective meansof targeting therapeutic doses of IL-2 to p185 positive tumors without increasing systemic toxicity or immunogenicity.

  14. Development of high-affinity single chain Fv against foot-and-mouth disease virus.

    Science.gov (United States)

    Jung, Joon-Goo; Jeong, Gu Min; Yim, Sung Sun; Jeong, Ki Jun

    2016-03-01

    Foot-and-mouth disease (FMD) is caused by the FMD virus (FMDV) and results in severe economic losses in livestock farming. For rapid FMD diagnostic and therapeutic purposes, an effective antibody against FMDV is needed. Here, we developed a high-affinity antibody against FMDV by FACS-based high throughput screening of a random library. With the FITC-conjugated VP1 epitope of FMDV and high-speed FACS sorting, we screened the synthetic antibody (scFv) library in which antibody variants are displayed in the periplasm of Escherichia coli. After three rounds of sorting, we isolated one antibody fragment (#138-scFv) against the VP1 epitope of FMDV. Next, to improve its affinity, a mutation library of #138-scFV was constructed by error-prone PCR and screened by FACS. After three rounds of sorting, we isolated one antibody (AM-32 scFv), which has a higher binding affinity (KD=42.7nM) than that of the original #138-scFv. We also confirmed that it specifically binds to whole inactivated FMDV. PMID:26827774

  15. Codon modification for the DNA sequence of a single-chain Fv antibody against clenbuterol and expression in Pichia pastoris

    Science.gov (United States)

    To improve expression efficiency of the recombinant single-chain variable fragment (scFv) against clenbuterol (CBL) obtained from mouse in the methylotrophic yeast Pichia pastoris (P. pastoris) GS115, the DNA sequence encoding for CBL-scFv was designed and synthesized based on the codon bias of P. p...

  16. Preparation of human single chain Fv antibody against hepatitis C virus E2 protein and its identification in immunohistochemistry

    Institute of Scientific and Technical Information of China (English)

    Yan-Wei Zhong; Jun Cheng; Gang Wang; Shuang-Shuang Shi; Li Li; Ling-Xia Zhang; Ju-Mei Chen

    2002-01-01

    AIM: To screen human single chain Fv antibody (scFv)against hepatitis C virus E2 antigen and identify its applicationin immunohistochemistry.METHODS: The phage antibody library was panned by HCVE2 antigen, which was coated in microtiter plate. After fiverounds of biopanning,56 phage clones were identified specificto HCV E2 antigen. The selected scFv clones were digestedby SfiI/NotI and DNA was sequenced. Then it was subclonedinto the vector pCANTABSE for expression as E-taggedsoluble scFv. The liver tissue sections from normal personand patients with chronic hepatitis B and chronic hepatitis Cwere immunostained with HCV E2 scFv antibody.RESULTS: The data of scFv-E2 DNA digestion and DNAsequencing showed that the scFv gene is composed of 750bp. ELISA and immunohistochemistry demonstrated that thehuman single chain Fy antibody against hepatitis C E2 antigenhas a specific binding character with hepatitis virus E2 antigenand paraffin-embedded tissue, but did not react with liver tissuesfrom healthy persons or patients with chronic hepatitis B.CONCLUSION: We have successfully screened andidentified HCV E2 scFv and the scFv could be used in theimmunostaining of liver tissue sections from patients withchronic hepatitis C.

  17. Soluble Expression, Purification and Characterization of Single-chain Fv Catalytic Antibody(sFv-2F3)

    Institute of Scientific and Technical Information of China (English)

    SUN Ye; LI Wei-jia; MA Ji-sheng; MU Ying; DU Xiu-bo; YAN Gang-lin; LUO Gui-min

    2004-01-01

    To find optimal conditions for expressing the soluble form of sFv-2F3 and to study the purification and property of its derivative Se-sFv-2F3, the preferred expression conditions were investigated by means of orthogonal design. These culture conditions included incubation temperature, inducer concentration, induction time and cell concentration. The evaluation of expression was accomplished by the analysis of whole cell lysates and the yield of soluble sFv-2F3 was calculated according to the analysis of Profinder(FTI-500,Pharmacia). The purification procedure was carried out via a two-step purification procedure consisting of ion-exchange chromatography, followed by immobilized metal affinity chromatography(IMAC). The antioxidant efficacy of Se-sFv-2F3 was demonstrated by the determination of the content of the main product of lipid peroxidation, MDA, the viability of cells and the activity of LDH. We obtained the preferred culture conditions to grow the engineered bacteria and the procedure for preparing soluble sFv-2F3 and confirmed the antioxidant efficacy of Se-sFv-2F3.

  18. Construction of scFv that bind both fibronectin-binding protein A and clumping factor A of Stapylococcus aureus.

    Science.gov (United States)

    Wang, Man; Zhang, Yan; Li, Benqiang; Zhu, Jianguo

    2015-06-01

    Bovine mastitis (BM) causes significant losses to the dairy industry. Vaccines against the causative agent of BM, Staphylococcus aureus, do not confer adequate protection. Because passive immunization with antibodies permits disease prevention, we constructed a recombinant single-chain antibody (scFv) against fibronectin-binding protein A (FnBPA) and clumping factor A (ClfA), two important virulence factors in S. aureus infection. The DNA coding sequences of the variable heavy (VH) and variable light (VL) domains of antibodies produced in the peripheral blood lymphocytes of cows with S. aureus-induced mastitis were obtained using reverse transcription and polymerase chain reaction, and the VH and VL cDNAs were assembled in-tandem using a DNA sequence encoding a (Gly4Ser)3 peptide linker. The scFv cDNAs were cloned into the pOPE101 plasmid for the expression of soluble scFv protein in Escherichia coli. The binding of the scFvs to both FnBPA and ClfA was confirmed using an indirect ELISA and Western blotting. The DNA sequences of the framework regions of the VH and VL domains were highly conserved, and the complementarity-determining regions displayed significant diversity, especially in CDR3 of the VH domain. These novel bovine antibody fragments may be useful as a therapeutic candidate for the prevention and treatment of S. aureus-induced bovine mastitis. PMID:25910693

  19. Characterization of a Single Chain Fv Antibody that Reacts with Free Morphine

    Directory of Open Access Journals (Sweden)

    Kazuhisa Sugimura

    2013-02-01

    Full Text Available An immune phage library derived from mice, hyperimmunized with morphine-conjugated BSA, was used to isolate a single-chain Fv (scFv clone, M86, with binding activity to morphine-conjugated thyroglobulin (morphine-C-Tg but not to codeine-, cocaine-, or ketamine-conjugated Tg. Surface plasmon resonance analysis using a morphine-C-Tg-coupled CM5 sensor chip showed that the Kd value was 1.26 × 10−8 M. To analyze its binding activity to free morphine and related compounds, we performed a competitive ELISA with M86 and morphine-C-Tg in the absence or presence of varying doses of free morphine and related compounds. IC50 values for opium, morphine, codeine, and heroin were 257 ng/mL, 36.4, 7.3, and 7.4 nM, respectively. Ketamine and cocaine exhibited no competitive binding activity to M86. Thus, we established a phage library-derived scFv, M86, which recognized not only free morphine and codeine as opium components but also heroin. This characteristic of M86 may be useful for developing therapeutic reagents for opiate addiction and as a free morphine-specific antibody probe.

  20. Generation and characterization of a human single-chain fragment variable (scFv antibody against cytosine deaminase from Yeast

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    Tombesi Marina

    2008-09-01

    Full Text Available Abstract Background The ability of cytosine deaminase (CD to convert the antifungal agent 5-fluorocytosine (5-FC into one of the most potent and largely used anticancer compound such as 5-fluorouracil (5-FU raised considerable interest in this enzyme to model gene or antibody – directed enzyme-prodrug therapy (GDEPT/ADEPT aiming to improve the therapeutic ratio (benefit versus toxic side-effects of cancer chemotherapy. The selection and characterization of a human monoclonal antibody in single chain fragment (scFv format represents a powerful reagent to allow in in vitro and in vivo detection of CD expression in GDEPT/ADEPT studies. Results An enzymatic active recombinant CD from yeast (yCD was expressed in E. coli system and used as antigen for biopanning approach of the large semi-synthetic ETH-2 antibody phage library. Several scFvs were isolated and specificity towards yCD was confirmed by Western blot and ELISA. Further, biochemical and functional investigations demonstrated that the binding of specific scFv with yCD did not interfere with the activity of the enzyme in converting 5-FC into 5-FU. Conclusion The construction of libraries of recombinant antibody fragments that are displayed on the surface of filamentous phage, and the selection of phage antibodies against target antigens, have become an important biotechnological tool in generating new monoclonal antibodies for research and clinical applications. The scFvH5 generated by this method is the first human antibody which is able to detect yCD in routinary laboratory techniques without interfering with its enzymatic function.

  1. Development of single-chain Fv against the nucleoprotein of type A influenza virus and its use in ELISA.

    Science.gov (United States)

    Sengupta, Devyani; Shaikh, Asma; Bhatia, S; Pateriya, A K; Khandia, R; Sood, R; Prakash, A; Pattnaik, B; Pradhan, H K

    2014-11-01

    Single chain fragment variable (ScFv) antibodies specific to the nucleoprotein (NP) of avian influenza virus (AIV) were developed using a phage display system. The variable heavy (VH) and the variable light (VL) chain gene fragments were derived from spleen cells of Balb/c mouse immunized with a recombinant NP (rNP) antigen (∼63 kDa) of H5N1 influenza virus. The VH and the VL DNA fragments were assembled through a flexible linker DNA to generate ScFv DNA that was cloned subsequently in a phagemid to express ScFv protein in Escherichia coli cells. The specific reactivity of the ScFv with the rNP antigen and viral antigen (H5N1) was confirmed by Western blot and ELISA. A competitive inhibition ELISA (CI-ELISA) was developed using the rNP and the anti-NP ScFv for detection of type-specific antibodies to AIV in chicken sera. The ScFv based CI-ELISA was compared with hemagglutination inhibition (HI) test and agar gel immunodiffusion (AGID) test over 850 sera. Sensitivity of the CI-ELISA was 100% with HI and AGID and specificity was 98.7% with HI and 100% with AGID. PMID:25152529

  2. Immunoreactivity and Radioimmunoscintigraphy of 4-Lysine Single Chain (Fv) Lym-1 Antibody for the Radiometal Chelation

    International Nuclear Information System (INIS)

    Small size of recombinant scFv, composed of VH and VL region of IgG, has many advantages such as faster blood clearance, improved tumor localization and reduced human anti-mouse antibody (HAMA) response. On the other hand, owing to small size, number of amino group, which was not involved in binding site, of ScFv lym-1 was insufficient in conjugation with CITC-DTPA chelator for radio metal labeling. The goal of this study is to introduce 4-lysine tag to the end of ScFv lym-1 sequence for radio metal conjugation and to evaluate the immunoreactivity and radioimmunoscintigraphy of chelator conjugated 4-lysine taq scFv lym-1 (4-lys scFv)

  3. Cloning approach and functional analysis of anti-intimin single-chain variable fragment (scFv

    Directory of Open Access Journals (Sweden)

    Elias Waldir P

    2011-02-01

    Full Text Available Abstract Background Intimin is an important virulence factor involved in the pathogenesis of enteropathogenic Escherichia coli (EPEC and enterohemorrhagic Escherichia coli (EHEC. Both pathogens are still important causes of diarrhea in children and adults in many developing and industrialized countries. Considering the fact that antibodies are important tools in the detection of various pathogens, an anti-intimin IgG2b monoclonal antibody was previously raised in immunized mice with the conserved sequence of the intimin molecule (int388-667. In immunoblotting assays, this monoclonal antibody showed excellent specificity. Despite good performance, the monoclonal antibody failed to detect some EPEC and EHEC isolates harboring variant amino acids within the 338-667 regions of intimin molecules. Consequently, motivated by its use for diagnosis purposes, in this study we aimed to the cloning and expression of the single-chain variable fragment from this monoclonal antibody (scFv. Findings Anti-intimin hybridoma mRNA was extracted and reversely transcripted to cDNA, and the light and heavy chains of the variable fragment of the antibody were amplified using commercial primers. The amplified chains were cloned into pGEM-T Easy vector. Specific primers were designed and used in an amplification and chain linkage strategy, obtaining the scFv, which in turn was cloned into pAE vector. E. coli BL21(DE3pLys strain was transformed with pAE scFv-intimin plasmid and subjected to induction of protein expression. Anti-intimin scFv, expressed as inclusion bodies (insoluble fraction, was denatured, purified and submitted to refolding. The protein yield was 1 mg protein per 100 mL of bacterial culture. To test the functionality of the scFv, ELISA and immunofluorescence assays were performed, showing that 275 ng of scFv reacted with 2 mg of purified intimin, resulting in an absorbance of 0.75 at 492 nm. The immunofluorescence assay showed a strong reactivity with

  4. An antigen-mediated selection system for mammalian cells that produce glycosylated single-chain Fv

    International Nuclear Information System (INIS)

    Selection and production of specific antibodies are limiting the development of high-throughput immunoassays such as antibody chips. In this study, we propose an antigen-mediated selection of antibody producers (ASAP) system in mammalian cells. As a model system, transgenes encoding anti-fluorescein ScFv fused to cytokine receptors were introduced to IL-3-dependent cell lines. Addition of fluorescein-conjugated BSA induced growth signal through the ScFv/receptor chimeras, leading to selective expansion of the transduced cells. Cre recombinase was then used to excise the receptor gene flanked by two loxP recognition sites in the introns, resulting in secretion of his-myc-tagged ScFv to the culture medium. When the first loxP site was used in the exon as a linker between ScFv and receptor, enhanced antigen-mediated cell proliferation and production of unexpectedly glycosylated ScFv were achieved. ASAP is the first mammalian selection/production system of recombinant human ScFvs, without need for subcloning and with the advantage of glycosylated product

  5. Development of single-chain variable fragments (scFv) against influenza virus targeting hemagglutinin subunit 2 (HA2).

    Science.gov (United States)

    Li, Tai-Wei; Cheng, Shu-Fang; Tseng, Yen-Tzu; Yang, Yu-Chih; Liu, Wen-Chun; Wang, Sheng-Cyuan; Chou, Mei-Ju; Lin, Yu-Jen; Wang, Yueh; Hsiao, Pei-Wen; Wu, Suh-Chin; Chang, Ding-Kwo

    2016-01-01

    Influenza A viruses (IAV) are widespread in birds and domestic poultry, occasionally causing severe epidemics in humans and posing health threats. Hence, the need to develop a strategy for prophylaxis or therapy, such as a broadly neutralizing antibody against IAV, is urgent. In this study, single-chain variable fragment (scFv) phage display technology was used to select scFv fragments recognizing influenza envelope proteins. The Tomlinson I and J scFv phage display libraries were screened against the recombinant HA2 protein (rHA2) for three rounds. Only the third-round elution sample of the Tomlinson J library showed high binding affinity to rHA2, from which three clones (3JA18, 3JA62, and 3JA78) were chosen for preparative-scale production as soluble antibody by E. coli. The clone 3JA18 was selected for further tests due to its broad affinity for influenza H1N1, H3N2 and H5N1. Simulations of the scFv 3JA18-HA trimer complex revealed that the complementarity-determining region of the variable heavy chain (VH-CDR2) bound the stem region of HA. Neutralization assays using a peptide derived from VH-CDR2 also supported the simulation model. Both the selected antibody and its derived peptide were shown to suppress infection with H5N1 and H1N1 viruses, but not H3N2 viruses. The results also suggested that the scFvs selected from rHA2 could have neutralizing activity by interfering with the function of the HA stem region during virus entry into target cells. PMID:26446888

  6. Development of single chain variable fragment (scFv) antibodies against Xylella fastidiosa subsp. pauca by phage display.

    Science.gov (United States)

    Yuan, Qing; Jordan, Ramon; Brlansky, Ronald H; Istomina, Olga; Hartung, John

    2015-10-01

    Xylella fastidiosa is a member of the gamma proteobacteria. It is fastidious, insect-vectored and xylem-limited and causes a variety of diseases, some severe, on a wide range of economically important perennial crops, including grape and citrus. Antibody based detection assays are commercially available for X. fastidiosa, and are effective at the species, but not at the subspecies level. We have made a library of scFv antibody fragments directed against X. fastidiosa subsp. pauca strain 9a5c (citrus) by using phage display technology. Antibody gene repertoires were PCR-amplified using 23 primers for the heavy chain variable region (V(H)) and 21 primers for the light chain variable region (V(L)). The V(H) and V(L) were joined by overlap extension PCR, and then the genes of the scFv library were ligated into the phage vector pKM19. The library contained 1.2×10(7) independent clones with full-length scFv inserts. In each of 3cycles of affinity-selection with 9a5c, about 1.0×10(12) phage were used for panning with 4.1×10(6), 7.1×10(6), 2.1×10(7) phage recovered after the first, second and third cycles, respectively. Sixty-six percent of clones from the final library bound X. fastidiosa 9a5c in an ELISA. Some of these scFv antibodies recognized strain 9a5c and did not recognize X. fastidiosa strains that cause Pierce's disease of grapevine. PMID:26232710

  7. Efficient construct of a large and functional scFv yeast display library derived from the ascites B cells of ovarian cancer patients by three-fragment transformation-associated recombination.

    Science.gov (United States)

    Yuan, Xiaopeng; Chen, Xiang; Yang, Mingjuan; Hu, Jia; Yang, Wei; Chen, Tingtao; Wang, Qirui; Zhang, Xuhua; Lin, Ruihe; Zhao, Aizhi

    2016-05-01

    Over the past decade, yeast display technology has emerged as a powerful tool for the isolation of high-affinity immunoglobulin fragments with potential utility as clinical diagnostic and therapeutic reagents. Despite significant refinement of the various methodologies underpinning library construction and selections, certain aspects remain challenging and process limiting. We have sought to significantly improve the robustness of the single-chain Fv (scFv) library construction step by overcoming the technical inefficiencies frequently encountered during the PCR-mediated assembly of scFvs from the discrete heavy and light V-domain repertoires. Using a novel primer set designed to provide maximum amplification coverage of the known germ-line V-domain repertoire, we have exploited the potential of the in vivo homologous gap-repair apparatus of Saccharomyces cerevisiae to assemble intact scFvs directly from co-transformed PBMC-derived VH, VL, and linearized vector component fragments. We have successfully applied this three-fragment assembly strategy to construct a large (>10(9)) scFv yeast display library from the ascites immune repertoire of ovarian cancer patients and validated the approach by applying FACS-based sorting to readily isolate scFvs that recognize various tumor marker antigens (TMAs). It is expected that this simplified construction method may find general utility, both for de novo scFv library construction and for subsequent combinatorial affinity maturation manipulations that require more than two fragments. PMID:26782745

  8. Construction and expression of D-dimer and GPIIb/IIIa single-chain bispecific antibody

    OpenAIRE

    DAN, ZHAOKUI; Tan, Zui; Xia, Hongli; WU, GAN

    2013-01-01

    The aim of this study was to construct a plasmid expressing glycoprotein IIb-IIIa (GPIIb/IIIa) and D-dimer single-chain bispecific antibody for the targeted therapy of thrombosis. The phosphorylated gene encoding the anti-GPIIb/IIIa single-chain variable fragment (scFv) and the gene encoding the anti-D-dimer scFv were amplified by PCR and linked in tandem by blunt-end ligation. The recombinant plasmid was transfected into the competent cell line HB2151 and identified by PCR and DNA sequencing...

  9. Construction of bifunctional molecules specific to antigen and antibody’s Fc-fragment by fusion of scFv-antibodies with staphylococcal protein A

    Directory of Open Access Journals (Sweden)

    Kolibo D. V.

    2009-06-01

    Full Text Available Aim. To develop approach for detection of scFv and their complexes with antigens. Methods. The fusion proteins, which include sequences of scFv and staphylococcal protein A, were constructed and the obtained bifunctional molecules were immunochemically analysed. Results. It was shown, that scFv fused with protein A and their complexes with antigens are effectively recognized by labelled immunoglobulins with unrestricted antigenic specificity. Conclusions. The fusion of scFv with protein A fragment is a perspective approach to increase the efficiency of application in ELISA. The obtained scFv, fused with protein A, could be used for development of test-systems for the detection of diphtheria toxin.

  10. Efficient refolding and immobilization of PMMA-tag-fused single-chain Fv antibodies for sensitive immunological detection on a PMMA plate.

    Science.gov (United States)

    Kumada, Yoichi; Ishikawa, Yasuyuki; Fujiwara, Yusuke; Takeda, Rui; Miyamoto, Ryosuke; Niwa, Daisuke; Momose, Shun; Kang, Bongmun; Kishimoto, Michimasa

    2014-09-01

    In this study, we investigated the efficient refolding and site-specific immobilization of single-chain variable fragments (scFvs) genetically fused with a poly(methylmethacrylate)-binding peptide (PMMA-tag). According to the results of an aggregation test of a scFv-PM in the presence of 0.5 M urea, aggregation was hardly detectable at a weak-alkaline pH (8.5) with lower concentrations of NaCl. Consequently, more than 93% recovery of the anti-RNase scFv-PM model was attained, when it was refolded by dialysis against 50 mM TAPS (pH8.5). These results suggested that the apparent isoelectric point (pI) of a target scFv was decreased to a great extent by the genetic fusion of a PMMA-tag containing 5 acidic amino acids, and, thus, the solubility of the scFv-PM in its semi-denatured form was considerably improved. We also designed alternative peptide-tags composed of plural aspartic acid residues (D5, D10 and D15-tags) to decrease the apparent pI value of the fusion protein. As a consequence, scFv-D5, scFv-D10 and scFv-D15 were also efficiently refolded with yields of more than 95%. It is noteworthy that even scFv-PS-D15, which had both a positively charged polystyrene-binding peptide (PS-tag) and a negatively charged D15-tag, was serially connected at the C-terminal region of scFvs, and also refolded with a yield of 96.1%. These results clearly indicate that controlling the apparent pI value of scFvs by the fusion of oligo-peptides composed of acidic amino acids at the C-terminus resulted in a high degree of recovery via dialysis refolding. According to the results of a sandwich ELISA using scFv-PMs, scFv-D15 and scFv-PS-D15 as ligands, high antigen-binding signals were detected from both the PMMA and phi-PS plates immobilized with scFv-PMs. Furthermore, the high antigen-binding activity of scFv-PMs was maintained in an adsorption state when it was immobilized on the surface of not only PMMA, but also hydrophilic PS (phi-PS) and polycarbonate (PC). These results

  11. Construction and bacterial expression of a recombinant single-chain antibody fragment against Wuchereria bancrofti SXP-1 antigen for the diagnosis of lymphatic filariasis.

    Science.gov (United States)

    Kamatchi, R; Charumathi, J; Ravishankaran, R; Kaliraj, P; Meenakshisundaram, S

    2016-01-01

    Global programmes to eliminate lymphatic filariasis (GPELF) require mapping, monitoring and evaluation using filarial antigen diagnostic kits. To meet this objective, a functional single-chain fragment variable (ScFv) specific for filarial Wuchereria bancrofti SXP-1 (Wb-SXP-1) antigen was constructed for the diagnosis of active filarial infection, an alternative to the production of complete antibodies using hybridomas. The variable heavy chain (VH) and the variable light chain (kappa) (Vκ) genes were amplified from the mouse hybridoma cell line and were linked together with a flexible linker by overlap extension polymerase chain reaction (PCR). The ScFv construct (Vκ-Linker-VH) was expressed as a fusion protein with N-terminal His tag in Escherichia coli and purified using immobilized metal affinity chromatography (IMAC) without the addition of reducing agents. Immunoblotting and sandwich enzyme-linked immunosorbent assay (ELISA) were used to analyse the antigen binding affinity of purified ScFv. The purified ScFv was found to recognize recombinant and native Wb-SXP-1 antigen in microfilariae (Mf)-positive patient sera. The affinity of ScFv was comparable with that of the monoclonal antibody. The development of recombinant ScFv to replace monoclonal antibody for detection of filarial antigen was achieved. The recombinant ScFv was purified, on-column refolded and its detection ability validated using field samples. PMID:26693887

  12. Construction of Large Human Single-chain Antibody Phage Display Library

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    A large human naive single chain antibody (scFv) library is constructed from 60 healthy donors via phage display technique. During the period, some methods are employed to optimize the diversity, such as multi donors, different annealing temperature, half-nest PCR, and assembly by two-way fusion PCR. In this stud y, 78 electroporations resulted in 1010 library, diversity of which is assayed by enzyme fingerprint. The efficiency and diversity are all better than other rese arches.

  13. Production and characterization of a recombinant single-chain antibody (scFv) for tracing the σ54 factor of Pseudomonas putida.

    Science.gov (United States)

    Jurado, Paola; Fernández, Luis Angel; de Lorenzo, Víctor

    2012-07-31

    The number of alternative sigma factor molecules per bacterial cell determines either stochasticity or evenness of transcription of cognate promoters. An approach for examining the abundance of sigmas in any sample of bacterial origin is explained here which relies on the production of a recombinant highly specific, high-affinity single-chain variable Fv domain (scFv) targeted towards unique protein sites of the factor. Purposely, a super-binder scFv recognizing a distinct epitope of the less abundant sigma σ(54) of Pseudomonas putida (also known as σ(N)) was obtained and its properties examined in detail. To this end, an scFv library was generated from mRNA extracted from lymphocytes of mice immunized with the purified σ(54) protein of this bacterium. The library was displayed on a phage system and subjected to various rounds of panning with purified σ(54) for capturing extreme binders. The resulting high-affinity anti-σ(54) phage antibody (Phab) clone named C2 strongly attached a small region located between positions 172 and 183 of the primary amino acid sequence of σ(54) that overlaps its core RNA polymerase-binding region. The purified scFv-C2 detected minute amounts of σ(54) in whole cell protein extracts not only of P. putida but also Escherichia coli cells and putatively in other bacteria as well. The affinity constant of the purified antibody was measured by surface plasmon resonance (SPR) and found to have a K(D) (k(off)/k(on)) in the range of 2×10(-9)M. The considerable affinity and specificity of this recombinant antibody makes it a tool of choice for quantitative studies on gene expression of σ(54)-dependent promoters in P. putida and other Gram-negative bacteria. PMID:22206981

  14. Development of single chain variable fragment (scFv) antibodies against surface proteins of 'Ca. Liberibacter asiaticus'.

    Science.gov (United States)

    Yuan, Qing; Jordan, Ramon; Brlansky, Ronald H; Minenkova, Olga; Hartung, John

    2016-03-01

    'Candidatus Liberibacter asiaticus' is the causal agent of citrus huanglongbing, the most serious disease of citrus worldwide. We have developed and applied immunization and affinity screening methods to develop a primary library of recombinant single chain variable fragment (scFv) antibodies in an M13 vector, pKM19. The antibody population is enriched for antibodies that bind antigens of 'Ca. Liberibacter asiaticus'. The primary library has more than 10(7) unique antibodies and the genes that encode them. We have screened this library for antibodies that bind to specifically-chosen proteins that are present on the surface of 'Ca. Liberibacter asiaticus'. These proteins were used as targets for affinity-based selection of scFvs that bind to the major outer membrane protein, OmpA; the polysaccharide capsule protein KpsF; a protein component of the type IV pilus (CapF); and, two flagellar proteins FlhA and FlgI. These scFvs have been used in ELISA and dot blot assays against purified protein antigens and 'Ca. Liberibacter asiaticus' infected plant extracts. We have also recloned many of these scFvs into a plasmid expression vector designed for the production of scFvs. Screening of these scFvs was more efficient when phage-bound, rather than soluble scFvs, were used. We have demonstrated a technology to produce antibodies at will and against any protein target encoded by 'Ca. Liberibacter asiaticus'. Applications could include advanced diagnostic methods for huanglongbing and the development of immune labeling reagents for in planta applications. PMID:26744234

  15. Targeted therapy against human lung cancer in nude mice by high-affinity recombinant antimesothelin single-chain Fv immunotoxin.

    Science.gov (United States)

    Fan, Dominic; Yano, Seiji; Shinohara, Hisashi; Solorzano, Carmen; Van Arsdall, Melissa; Bucana, Corazon D; Pathak, Sen; Kruzel, Ewa; Herbst, Roy S; Onn, Amir; Roach, Jennifer S; Onda, Masanori; Wang, Qing-cheng; Pastan, Ira; Fidler, Isaiah J

    2002-06-01

    Several tumors, including mesothelioma and ovarian cancer, can overexpress mesothelin, a glycosylphosphatidylinositol-linked differentiation glycoprotein. The membrane-bound type of mesothelin is found in the blood of cancer patients at a very low level, which makes mesothelin a good candidate for targeted therapy of certain cancers. An antimesothelin disulfide-linked Fv (SS1 Fv) was fused to a truncated mutant of Pseudomonas exotoxin A to produce the recombinant immunotoxin SS1(dsFv)-PE38, which has a high binding affinity to mesothelin (Kd = 0.7 nM). Our studies in vitro showed that SS1(dsFv)-PE38 is significantly more cytotoxic to the high-mesothelin-producing NCI-H226 human non-small cell lung cancer cells than to human lung adenocarcinoma PC14PE6 cells, which do not express mesothelin. When administered at a nontoxic dose of 500 microg/kg on days 7, 9, and 11 to nude mice injected i.v. with the two human lung cancer cell lines, SS1(dsFv)-PE38 selectively inhibited experimental lung metastases produced by the mesothelin-producing NCI-H226 cells. Our data indicate that mesothelin-producing squamous cell carcinoma of the lung may be a good target for this immunotoxin. PMID:12479219

  16. [Expression purification and verification of HBscFv-IFNgamma in Pichia pastoris x33].

    Science.gov (United States)

    Zhou, Shishui; Wang, Xiaoning

    2008-03-01

    In order to effectively cure hepatitis B virus (HBV), we studied on fusion protein HBscFv-IFNgamma, which was connected with single-chain Fv against HBV surface antigen(HBscFv) and gamma-interferon(IFNgamma) of being used in clinic against HBV. Adopting overlap PCR, the hbscfv and the ifngamma were connected into hbscfv-ifngamma. Then the pPICZalphaA/(hbscfv-ifngamma)(1,2,4) of multi-copy recombinant plasmid were constructed and transformed into Pichia pastoris x33. The engineering strain x4 was screened from transformed x33 and could secretively express HBscFv-IFNgamma. The preliminary verification indicates that HBscFv-IFNgamma has the bioactivity of HBscFv and IFNgamma by SDS-PAGE, Western blotting and ELISA. The supernatant of culturing X4 was purified by 14F7 affinity chromatography to HBscFv-IFNgamma with purity of 95%-98%. The HBscFv-IFNgamma is able to bind 27.9% HBV surface antigen (HBsAg) in the serum of HBV transgenic mice, which shows the antibody of HBscFv-IFNgamma has bioactivity in vivo. Therefore HBscFv-IFNgamma can shed light on the development of a new promising HBV-targeted drug. PMID:18589818

  17. Improving construction performance through supply chain management

    Institute of Scientific and Technical Information of China (English)

    王要武; 薛小龙

    2004-01-01

    A typical model of construction supply chain (CSC) is presented firstly. Then the characteristics and problems of CSC are identified. The authors also describe the progress towards adoption of supply chain management (SCM) in the construction industry. Traditional construction management approaches (CMAs) are compared with new SCM-based CMAs. The authors suggest that integrated SCM based on cooperating competition is crucial to improve construction performance. Based on relevant results from related research, a framework for integrated construction supply chain management (CSCM) is developed to guide the effective implementation of SCM in construction.

  18. Understanding Construction Supply Chains: An Alternative Interpretation

    OpenAIRE

    Vrijhoef, Ruben; Koskela, Lauri; Howell, Greg

    2001-01-01

    Much research work has assessed that construction is ineffective and many problems can be observed. Analysis of these problems has shown that a major part of them are supply chain problems, originating at the interfaces of different parties or functions. There have been several kinds of initiatives aiming at improvement and renewal of construction supply chains, but only few have a track record of consequent and significant successes. Here construction supply chains are approached from an alt...

  19. Application of adaptive DO-stat feeding control to Pichia pastoris X33 cultures expressing a single chain antibody fragment (scFv).

    Science.gov (United States)

    Ferreira, A R; Ataíde, F; von Stosch, M; Dias, J M L; Clemente, J J; Cunha, A E; Oliveira, R

    2012-11-01

    In this study, fed-batch cultures of a Pichia pastoris strain constitutively expressing a single chain antibody fragment (scFv) under the control of the glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter were performed in a pilot 50 L bioreactor. Due to the very high cell density achieved within the first 75 h, typically between 140 and 160 g-DCW/L of dry cell weight (DCW), most of the scFv is produced under hard oxygen transfer limitation. To improve scFv productivity, a direct adaptive dissolved oxygen (DO)-stat feeding controller that maximizes glycerol feeding under the constraint of available oxygen transfer capacity was developed and applied to this process. The developed adaptive controller enabled to maximize glycerol feeding through the regulation of DO concentration between 3 and 5 % of saturation, thereby improving process productivity. Set-point convergence dynamics are characterized by a fast response upon large perturbations to DO, followed by a slower but very robust convergence in the vicinity of the boundary with almost imperceptible overshoot. Such control performance enabled operating closer to the 0 % boundary for longer periods of time when compared to a traditional proportional-integral-derivative algorithm, which tends to destabilize with increasing cell density. PMID:22610694

  20. Expression of anti-tumor necrosis factor alpha (TNFα) single-chain variable fragment (scFv) in Spirodela punctata plants transformed with Agrobacterium tumefaciens.

    Science.gov (United States)

    Balaji, Parthasarathy; Satheeshkumar, P K; Venkataraman, Krishnan; Vijayalakshmi, M A

    2016-05-01

    Therapeutic antibodies against tumor necrosis factor alpha (TNFα) have been considered effective for some of the autoimmune diseases such as rheumatoid arthritis, Crohn's diseases, and so on. But associated limitations of the current therapeutics in terms of cost, availability, and immunogenicity have necessitated the need for alternative candidates. Single-chain variable fragment (scFv) can negate the limitations tagged with the anti-TNFα therapeutics to a greater extent. In the present study, Spirodela punctata plants were transformed with anti-TNFα through in planta transformation using Agrobacterium tumefaciens strain, EHA105. Instead of cefotaxime, garlic extract (1 mg/mL) was used to remove the agrobacterial cells after cocultivation. To the best of our knowledge, this report shows for the first time the application of plant extracts in transgenic plant development. 95% of the plants survived screening under hygromycin. ScFv cDNA integration in the plant genomic DNA was confirmed at the molecular level by PCR. The transgenic protein expression was followed up to 10 months. Expression of scFv was confirmed by immunodot blot. Protein expression levels of up to 6.3% of total soluble protein were observed. β-Glucuronidase and green fluorescent protein expressions were also detected in the antibiotic resistant plants. The paper shows the generation of transgenic Spirodela punctuata plants through in planta transformation. PMID:25786575

  1. Charge-modified single chain antibody constructs of monoclonal antibody CC49: generation, characterization, pharmacokinetics, and biodistribution analysis

    International Nuclear Information System (INIS)

    A novel strategy was developed in which an antibody scFv fragment of the monoclonal antibody (MAb) CC49 was modified by engineering DNA coding sequences to lower its isoelectric point. Negatively charged amino acids were added to the carboxy terminus of the CC49 VH region by adding nucleotide sequences in a polymerase chain reaction (PCR) amplification of the coding sequence of CC49 scFv. Two new DNA constructs coding for CC49 scFv with lower isoelectric points of 5.8 and 5.2 were engineered. These novel strategy-generated, charge-modified antibody constructs were compared for their immunological, pharmacokinetic, and biodistribution properties in athymic mice bearing LS-174T human colon carcinoma xenografts

  2. Generation and functional characterization of the anti-transferrin receptor single-chain antibody-GAL4 (TfRscFv-GAL4 fusion protein

    Directory of Open Access Journals (Sweden)

    Ye Qing

    2012-11-01

    Full Text Available Abstract Background The development of vectors for cell-specific gene delivery is a major goal of gene therapeutic strategies. Transferrin receptor (TfR is an endocytic receptor and identified as tumor relative specific due to its overexpression on most tumor cells or tissues, and TfR binds and intakes of transferrin-iron complex. We have previously generated an anti-TfR single-chain variable fragments of immunoglobulin (scFv which were cloned from hybridoma cell line producing antibody against TfR linked with a 20 aa-long linker sequence (G4S4. In the present study, the anti-TfR single-chain antibody (TfRscFv was fused to DNA-binding domain of the yeast transcription factor GAL4. The recombinant fusion protein, designated as TfRscFv-GAL4, is expected to mediate the entry of DNA-protein complex into targeted tumor cells. Results Fusion protein TfRscFv-GAL4 was expressed in an E. coli bacterial expression system and was recovered from inclusion bodies with subsequent purification by metal-chelate chromatography. The resulting proteins were predominantly monomeric and, upon refolding, became a soluble biologically active bifunctional protein. In biological assays, the antigen-binding activity of the re-natured protein, TfRscFv-GAL4, was confirmed by specific binding to different cancer cells and tumor tissues. The cell binding rates, as indicated by flow cytometry (FCM analysis, ranged from 54.11% to 8.23% in seven different human carcinoma cell lines. It showed similar affinity and binding potency as those of parent full-length mouse anti-TfR antibody. The positive binding rates to tumor tissues by tissue microarrays (TMA assays were 75.32% and 63.25%, but it showed weakly binding with hepatic tissue in 5 cases, and normal tissues such as heart, spleen, adrenal cortex blood vessel and stomach. In addition, the re-natured fusion protein TfRscFv-GAL4 was used in an ELISA with rabbit anti-GAL4 antibody. The GAL4-DNA functional assay through the GAL4

  3. Supply Chain Management in the Construction Industry

    OpenAIRE

    Olsson, Fredrik

    2000-01-01

    In this licentiate thesis the implications of a deliberate use of logistics knowledge have been investigated by describing and analyzing the use of a supply chain management approach in the construction industry.
    A five-step approach is used in the thesis. First, theoretical models are developed from the supply chain management literature. Two models are elaborated; One model identifying supply chain criteria and the other comparing traditional to supply chain-based.

  4. A large semi-synthetic single-chain Fv phage display library based on chicken immunoglobulin genes

    Directory of Open Access Journals (Sweden)

    Jordaan Frances

    2004-04-01

    Full Text Available Abstract Background Antibody fragments selected from large combinatorial libraries have numerous applications in diagnosis and therapy. Most existing antibody repertoires are derived from human immunoglobulin genes. Genes from other species can, however, also be used. Because of the way in which gene conversion introduces diversity, the naïve antibody repertoire of the chicken can easily be accessed using only two sets of primers. Results With in vitro diagnostic applications in mind, we have constructed a large library of recombinant filamentous bacteriophages displaying single chain antibody fragments derived from combinatorial pairings of chicken variable heavy and light chains. Synthetically randomised complementarity determining regions are included in some of the heavy chains. Single chain antibody fragments that recognise haptens, proteins and virus particles were selected from this repertoire. Affinities of three different antibody fragments were determined using surface plasmon resonance. Two were in the low nanomolar and one in the subnanomolar range. To illustrate the practical value of antibodies from the library, phage displayed single chain fragments were incorporated into ELISAs aimed at detecting African horsesickness and bluetongue virus particles. Virus antibodies were detected in a competitive ELISA. Conclusion The chicken-derived phage library described here is expected to be a versatile source of recombinant antibody fragments directed against a wide variety of antigens. It has the potential to provide monoclonal reagents with applications in research and diagnostics. For in vitro applications, naïve phage libraries based on avian donors may prove to be useful adjuncts to the selectable antibody repertoires that already exist.

  5. Achieving Supply Chain Integration within Construction Industry

    OpenAIRE

    Peter McDermotti; Malik Khalfan

    2012-01-01

    The main driver behind the adoption of supply chain management (SCM) philosophy into the construction industry was the successes within other industry sectors. SCM can be defined as network of different organisations, linked upstream and downstream in a chain, aiming to produce quality and value in the services and products for the end consumers through integrated processes and activities. In order to achieve the optimised level of integration of the whole supply chain, the industry has respo...

  6. Preparation and Identification of a Human Single Chain Fv Antibody Against Ascaris lumbricoides%抗蛔虫人源单链抗体库的构建及鉴定

    Institute of Scientific and Technical Information of China (English)

    何光志; 田维毅; 高英; 王平; 王文佳; 奚锦; 俞琦; 王乾宇; 黄高

    2011-01-01

    Objective To construct humanize phage antibody library against Ascaris lumbricoides and to screen specificity scFv to Ascaris lumbricoides. Methods Total RNA was abstracted from peripheral blood lympho-cytes of 20 persons, and was used to amplify VH and VL gene by RT-PCR. The amplified VH and VL genes were spliced to form scFv gene which was cloned into pCANTAB-SE phagemid, and the constructed recombinant phage-mid was transformed to E. Coli TC1 to construct human natural single-chain antibody library from which positive clones were screened. Results A primary library of 1.5 × 106 and a second library of 1.2 × 106 were constructed. Conclusion This study was to provide us the basis for radionuclide imaging and therapy for ascariasis.%目的:构建抗蛔虫人源单链抗体库,从中筛选建抗蛔虫人源特异性单链抗体.方法:分离10个患蛔虫病人的淋巴细胞,提取总RNA反转录为cDNA,PCR扩增人抗体重链(VH)和轻链(VL)可变区基因,采用SOE-PCR法将VH和VL片段随机拼接成scFv片段,并克隆入噬菌粒载体pCANTAB5E中,构建噬菌体单链抗体库.结果:初级库库容量为1.8×106,在大肠杆菌TG1中重组后得到1.6×106的次级抗体库.结论:本研究成功构建抗蛔虫人源单链抗体库,拟在为蛔虫病的预防、诊断、治疗奠定基础.

  7. Genetically engineered multivalent single chain antibody constructs for cancer therapy

    International Nuclear Information System (INIS)

    increase its tumor: normal tissue ratio for improved therapeutic index, we engineered a variety antibody constructs. These constructs were evaluated using novel approaches like special radionuclides, pretargeting and optimization. Due to the smaller size, the engineered antibody molecules should penetrate better throughout a tumor mass, with less dose heterogeneity, than is the case with intact IgG. Multivalent scFvs with an appropriate radionuclide, therefore, hold promising prospects for cancer therapy and clinical imaging in MAb-based radiopharmaceuticals. In addition, the human anti-mouse antibodies (HAMA) responses in patients against antibody-based therapy are usually directed against the immunoglobulin constant regions; however, anti-idiotypic responses can also be detected. The HAMA responses reduce the efficacy of treatment by removing the circulating antibody molecules, fragments, and possibly scFvs by altering the pharmacokinetic properties of the antibody. HAMA responses against divalent IgG, divalent Ig fragments, and possibly multimeric scFvs could cause immune complex formation with hypersensitivity or allergic reactions that could be harmful to patients. The use of small molecules, such as scFvs (monomeric as well as multimeric), with their shorter biological half-lives and the lack of the constant regions and humanized variable (binding regions) performed in our studies should reduce the development of HAMA. The generation of humanized and fully human scFvs should further reduce the development of HAMA. Specific accomplishments on the project are the production of large amounts of recombinant antibodies as they are required in large amounts for cancer diagnosis and therapy. A variety of single-chain Fv (scFv) constructs were engineered for the desired pharmacokinetic properties. Tetrameric and dimeric scFvs showed a two-fold advantage: (1) there was a considerable gain in avidity as compared to smaller fragments, and (2) the biological half-life was more

  8. Genetically engineered multivalent single chain antibody constructs for cancer therapy

    Energy Technology Data Exchange (ETDEWEB)

    Surinder Batra, Ph D

    2006-02-27

    its tumor: normal tissue ratio for improved therapeutic index, we engineered a variety antibody constructs. These constructs were evaluated using novel approaches like special radionuclides, pretargeting and optimization. Due to the smaller size, the engineered antibody molecules should penetrate better throughout a tumor mass, with less dose heterogeneity, than is the case with intact IgG. Multivalent scFvs with an appropriate radionuclide, therefore, hold promising prospects for cancer therapy and clinical imaging in MAb-based radiopharmaceuticals. In addition, the human anti-mouse antibodies (HAMA) responses in patients against antibody-based therapy are usually directed against the immunoglobulin constant regions; however, anti-idiotypic responses can also be detected. The HAMA responses reduce the efficacy of treatment by removing the circulating antibody molecules, fragments, and possibly scFvs by altering the pharmacokinetic properties of the antibody. HAMA responses against divalent IgG, divalent Ig fragments, and possibly multimeric scFvs could cause immune complex formation with hypersensitivity or allergic reactions that could be harmful to patients. The use of small molecules, such as scFvs (monomeric as well as multimeric), with their shorter biological half-lives and the lack of the constant regions and humanized variable (binding regions) performed in our studies should reduce the development of HAMA. The generation of humanized and fully human scFvs should further reduce the development of HAMA. Specific accomplishments on the project are the production of large amounts of recombinant antibodies as they are required in large amounts for cancer diagnosis and therapy. A variety of single-chain Fv (scFv) constructs were engineered for the desired pharmacokinetic properties. Tetrameric and dimeric scFvs showed a two-fold advantage: (1) there was a considerable gain in avidity as compared to smaller fragments, and (2) the biological half-life was more

  9. Production of recombinant single chain antibodies (scFv) in vegetatively reproductive Kalanchoe pinnata by in planta transformation.

    Science.gov (United States)

    Jung, Yuchul; Rhee, Yong; Auh, Chung-Kyoon; Shim, Hyekyung; Choi, Jung-Jin; Kwon, Suk-Tae; Yang, Joo-Sung; Kim, Donggiun; Kwon, Myung-Hee; Kim, Yong-Sung; Lee, Sukchan

    2009-10-01

    We developed an asexual reproductive plant, Kalanchoe pinnata, as a new bioreactor for plant-based molecular farming using a newly developed transformation method. Leaf crenate margins were pin-pricked to infect the plant with the Agrobacterium strain LBA4404 and vacuum infiltration was also applied to introduce the target gene into the plants. Subsequently, the young mother leaf produced new clones at the leaf crenate margins without the need for time- and labor-consuming tissue culture procedures. The average transformation rates were approximately 77 and 84% for pin-prickling and vacuum-infiltration methods, respectively. To functionally characterize an introduced target protein, a nucleic acid hydrolyzing recombinant 3D8 scFv was selected and the plant based 3D8 scFv proteins were purified and analyzed. Based on abzyme analysis, the purified protein expressed with this system had catalytic activity and exhibited all of properties of the protein produced in an E. coli system. This result suggested that vegetatively reproductive K. pinnata can be a novel and potent bioreactor for bio-pharmaceutical proteins. PMID:19688214

  10. Quality in Construction- A Supply Chain Perspective

    DEFF Research Database (Denmark)

    Koch, Christian; Larsen, Casper Schultz

    2006-01-01

    transformed into the material stream. The paper proposes initiatives to strengthen partnerships in supply chains, especially at mixed stable and project configured types. The contradiction between permanent enterprise organisations potentially capable of handling purchasing and the role of the project manager......Often the construction supply chain is perceived as consisting of two intertwined streams; a stream of materials and a stream of immaterial informational character. Moreover the supply delivery would be characterized by configuration by project. That this configuration is not always successful can...

  11. Humoral immune responses induced by anti-idiotypic antibody fusion protein of 6B11scFv/hGM-CSF in BALB/c mice

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Background We have previously developed and characterized a monoclonal anti-idiotype antibody, designated 6B11, which mimics an ovarian carcinoma associated antigen OC166-9 and whose corresponding monoclonal antibody is COC166-9 (Ab1). In this study, we evaluate the humoral immune responses induced by the fusion protein 6B11 single-chain variable fragment (scFv)/human granulocyte macrophage colony-stimulating factor (hGM-CSF) and 6B11scFv in BALB/c mice. Methods The fusion protein 6B11scFv/hGM-CSF was constructed by fusing a recombinant single-chain variable fragment of 6B11scFv to GM-CSF. BALB/c mice were administrated by 6B11scFv/hGM-CSF and 6B11scFv, respectively. Results The fusion protein 6B11scFv/hGM-CSF retained binding to the anti-mouse F(ab)2' and was also biologically active as measured by proliferation of human GM-CSF dependent cell TF1 in vitro. After immunization with the 6B11scFv/hGM-CSF and 6B11ScFv, BALB/c mice showed significantly enhanced Ab3 antibody responses to 6B11scFv/hGM-CSF compared with the 6B11scFv alone. The level of Ab3 was the highest after the first week and maintained for five weeks after the last immunization. Another booster was given when the Ab3 titer descended, and it would reach to the high level in a week. Conclusion The fusion protein 6B11scFv/hGM-CSF can induce humoral immunity against ovarian carcinoma in vivo. We also provide the theoretical foundation for the application of the fusion protein 6B11scFv/hGM-CSF for active immunotherapy of ovarian cancer.

  12. The use of a cocktail of single chain Fv antibody fragments to improve the in vitro and in vivo targeting of melanoma

    International Nuclear Information System (INIS)

    Radio scintigraphy using single chain antibody fragments (scFvs) offers a potenti al means of early detection of melanoma metastases. However, previous studies have shown suboptimal levels of tumour localization and nonspecific background accumulation which may be due to antigen heterogeneity. We aimed to improve tumour localization by using a cocktail of different scFvs targeting different epitopes on melanoma cells. We have previously developed three scFvs against distinct and highly tumour-specific melanoma cell-surface antigens by chain shuffling and antibody phage selection on melanoma cells. Three scFvs, RAFT3, B3 and B4 were labeled with 125Iodine and tested both individually and as a cocktail in a nude mouse xenograft model far human melanoma. Results demonstrated improved tumour localization in vivo when compared to the individual scFvs. Tumour uptake of the cocktail at l hour was 24.220% ID/g (injected dose/gram) compared with 2.854%, 2.263% and 1.355% far B4, RAFT3 and B3, respectively, when injected individually. In addition, the cocktail exhibited significantly superior tumour to normal tissue ratios far muscle and spleen (p<0.05). A combination or cocktail of scFv clones may have an advantage aver individual scFvs far melanoma targeting in patients because of heterogeneity in the expression of different epitopes of antigens on melanoma cells

  13. Characterization of an engineered human purine nucleoside phosphorylase fused to an anti-her2/neu single chain Fv for use in ADEPT

    Directory of Open Access Journals (Sweden)

    Wu Anna M

    2009-12-01

    Full Text Available Abstract Background Antibody Directed Enzyme Prodrug Therapy (ADEPT can be used to generate cytotoxic agents at the tumor site. To date non-human enzymes have mainly been utilized in ADEPT. However, these non-human enzymes are immunogenic limiting the number of times that ADEPT can be administered. To overcome the problem of immunogenicity, a fully human enzyme, capable of converting a non-toxic prodrug to cytotoxic drug was developed and joined to a human tumor specific scFv yielding a fully human targeting agent. Methods A double mutant of human purine nucleoside phosphorylase (hDM was developed which unlike the human enzyme can cleave adenosine-based prodrugs. For tumor-specific targeting, hDM was fused to the human anti-HER2/neu single chain Fv (scFv, C6 MH3B1. Enzymatic activity of hDM with its natural substrates and prodrugs was determined using spectrophotomeric approaches. A cell proliferation assay was used to assess the cytotoxicity generated following conversion of prodrug to drug as a result of enzymatic activity of hDM. Affinity of the targeting scFv, C6 MH3B1 fused to hDM to Her2/neu was confirmed using affinity chromatography, surface plasmon resonance, and flow-cytometry. Results In vitro hDM-C6 MH3B1 binds specifically to HER2/neu expressing tumor cells and localizes hDM to tumor cells, where the enzymatic activity of hDM-C6 MH3B1, but not the wild type enzyme, results in phosphorolysis of the prodrug, 2-fluoro-2'-deoxyadenosine to the cytotoxic drug 2-fluoroadenine (F-Ade causing inhibition of tumor cell proliferation. Significantly, the toxic small drug diffuses through the cell membrane of HER2/neu expressing cells as well as cells that lack the expression of HER2/neu, causing a bystander effect. F-Ade is toxic to cells irrespective of their growth rate; therefore, both the slowly dividing tumor cells and the non-dividing neighboring stromal cells that support tumor growth should be killed. Analysis of potential novel MHCII

  14. Production and Evaluation of Immunoreactivity of Poly Lysine-Tagged Single Chain Fragment Variable (ScFv) Lym-1 Antibody for Direct Conjugation to Fluorescence Dye

    International Nuclear Information System (INIS)

    Small size of recombinant scFv antibody has many advantages such as rapid blood clearances and improved targeting antibodies to tumor region. On the other hand owing to small size, number of amino group is insufficient in conjugation with chelator and fluorescence labeling. This study is to introduce poly lysine tag to the C-terminal end of scFv lym-1 sequence for fluorescence chelator conjugation. Poly lysine scFv lym-1 gene, cloned into pET-22b (+) vector, was expressed in E. coli BL21 (DE3) strain. Antibody purification was performed with Ni-NTA column and then size exclusion column chromatography. Expression and purification levels of poly lysine tagged scFv lym-1 antibody were confirmed by western blot analysis. I-124, I-125, I-131 and Tc-99m were used for radiolabeling of purified poly lysine scFv lym-1. Flow cytometry analysis of FITC conjugated poly lysine scFv lym-1 was performed for confirmation of immunoreactivity of human Burkitt's lymphoma cells. Poly lysine scFv lym-1 antibody was purified through two steps and identified as molecular weight of 48 KDa. Radiolabeling yields of I-124, I-125, I-131 and Tc-99m into poly lysine scFv lym-1 were >99%, >99%, >95% and >99%, respectively. Flow cytometry analysis of poly lysine scFv and scFv lym-1 was showed similar immunoreactivity to human Burkitt's lymphoma cells. Poly lysine tag was useful for the sufficient number of amino groups to scFv lym-1 antibody for chelator conjugation with minimizing loss of immunoreactivity

  15. Production and Evaluation of Immunoreactivity of Poly Lysine-Tagged Single Chain Fragment Variable (ScFv) Lym-1 Antibody for Direct Conjugation to Fluorescence Dye

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Jae Ho; Choi, Tae Hyun; Woo, Kwang Sun; Chung, Wee Sup; Kang, Joo Hyun; Jeong, Su Young; Choi, Chang Woon; Lim, Sang Moo; Cheon, Gi Jeong [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2009-10-15

    Small size of recombinant scFv antibody has many advantages such as rapid blood clearances and improved targeting antibodies to tumor region. On the other hand owing to small size, number of amino group is insufficient in conjugation with chelator and fluorescence labeling. This study is to introduce poly lysine tag to the C-terminal end of scFv lym-1 sequence for fluorescence chelator conjugation. Poly lysine scFv lym-1 gene, cloned into pET-22b (+) vector, was expressed in E. coli BL21 (DE3) strain. Antibody purification was performed with Ni-NTA column and then size exclusion column chromatography. Expression and purification levels of poly lysine tagged scFv lym-1 antibody were confirmed by western blot analysis. I-124, I-125, I-131 and Tc-99m were used for radiolabeling of purified poly lysine scFv lym-1. Flow cytometry analysis of FITC conjugated poly lysine scFv lym-1 was performed for confirmation of immunoreactivity of human Burkitt's lymphoma cells. Poly lysine scFv lym-1 antibody was purified through two steps and identified as molecular weight of 48 KDa. Radiolabeling yields of I-124, I-125, I-131 and Tc-99m into poly lysine scFv lym-1 were >99%, >99%, >95% and >99%, respectively. Flow cytometry analysis of poly lysine scFv and scFv lym-1 was showed similar immunoreactivity to human Burkitt's lymphoma cells. Poly lysine tag was useful for the sufficient number of amino groups to scFv lym-1 antibody for chelator conjugation with minimizing loss of immunoreactivity

  16. Inhibition of Breast Cancer Metastasis and Angiogenesis by Antiosteopontin Single-Chain Fv-Fc Fusion Protein

    Directory of Open Access Journals (Sweden)

    Ling Peng

    2009-05-01

    Full Text Available Osteopontin (OPN is associated with many diseases, and its role in tumor growth and metastasis has been studied in breast cancers. Previous studies have described anti-OPN antibodies that could inhibit tumor cell adhesion and invasion in vitro, but until now, there are no systematic studies on antitumor effects of anti-OPN antibodies in vivo. In the present study, we have raised several anti-OPN single-chain variable fragments from phage antibody library and expressed them as single-chain variable fragment-constant region fragment fusion proteins in Chinese hamster ovary cells. Of them, two antibodies (1A12 and 2H8 were able to inhibit MDA-MB-435s breast cancer cell attachment, invasion, migration, and colony formation in soft agar. Furthermore, 1A12 and 2H8 inhibited the anti-apoptotic and prosurvival functions of OPN in human umbilical vein endothelial cell. In human umbilical vein endothelial cell capillary tube formation, chicken chorioallantoic membrane assay, and rabbit corneal micropocket assay, the two antibodies showed markedly inhibitory effects toward angiogenesis. We investigated antitumor effects of anti-OPN antibodies in nude mice by assessing xenograft tumor growth and lung metastasis potential. The results showed that the two antibodies were capable of delaying primary tumor growth and reducing spontaneous lung metastasis. Epitope mappings of these two anti-OPN antibodies were performed, and a new binding site of 1A12 was revealed. In summary, the present study has demonstrated the roles of anti-OPN antibodies in blocking the function of OPN, suggesting that they may have the potential to be developed for future clinical use.

  17. 抗黄曲霉毒素单链抗体基因的克隆与表达%Cloning and expression of single chain Fv antibody gene against aflatoxin

    Institute of Scientific and Technical Information of China (English)

    李鑫; 李培武; 张奇; 张文; 李园园

    2012-01-01

    The objective of this study was to reduce the preparation cost of aflatoxin antibody. Based on the hybridoma cell 8F6 w0hich secrete mAbs against aflatoxin, the variable heavy (VH) and light ( VL) region genes of 8F6 were amplified using polymerase chain reaction. The VH - linker - VL genes were constructed by overlap extension, then cloned into a phagemid vector pCANTAB 5E and expressed in E. coli TG1 cells as a fusion with a phage protein. Indirect competitive ELISA was performed for detection of the binding activity, which resulted in IC50 value of the constructed ScFv as 0. 57 ng per milliliter.%为降低黄曲霉毒素抗体制备成本,在已有的能分泌抗黄曲霉毒素单克隆抗体的杂交瘤细胞系8F6的基础上,成功克隆得到了该单克隆抗体的重链(VH)和轻链可变区(VL)基因片段.通过重叠延伸PCR的方法将轻、重链可变区基因连接,并引入连接肽(Linker)编码序列,构建VH-Linker-VL结构的单链抗体(ScFv)基因,并将该基因克隆到噬菌体表达载体pCANTAB 5E上,使单链抗体以噬菌体展示形式在大肠杆菌TG1中表达.间接竞争ELISA方法检测到该ScFv对黄曲霉毒素B1的抑制率(IC50)值为0.57ng/mL,表明该单链抗体与亲本鼠单抗有相同的抗原结合特异性,且具有很高灵敏度.

  18. [Stable expression of human anti-IL-33 scFv-IgG1Fc fusion protein in CHO k1 cells].

    Science.gov (United States)

    Ye, Yingchun; Nian, Siji; Wang, Xu; Wu, Tong; Xu, Wenfeng; Yuan, Qing

    2016-05-01

    Objective To construct two different eukaryotic expression vectors of human anti-interleukin 33 (IL-33) single-chain antibody fragment (scFv-Fc) to transfect Chinese hamster ovary (CHO) k1 cells and select the stably and high-level expressed cell lines to improve the expression level of the fusion protein. Methods The previously constructed recombinant plasmid pcDNA3.1/SP-scFv-Fc was digested to obtain SP-scFv-Fc fragments, and the fragments were inserted into the plasmid PMH3(EN) to construct recombinant plasmid PMH3(EN)/SP-scFv-Fc. The plasmids PMH3(EN)/SP-scFv-Fc and pcDNA3.1/SP-scFv-Fc were separately transfected into CHO k1 cells. The transcription and translation level of the SP-scFv-Fc were detected by reverse transcription PCR (RT-PCR) and Western blotting, respectively. The stably and high-level expressed cell lines were screened by Dot blotting. The expression level and binding activity of the expressed scFv-Fc were measured by ELISA. Results The recombinant plasmid PMH3(EN)/SP-scFv-Fc was successfully constructed and the size of the inserted SP-scFv-Fc was about 1560 bp. The RT-PCR results showed that the SP-scFv-Fc was successfully transfected into CHO k1 cells. The scFv-Fc proteins could be secreted into the cultural supernatant and specifically bind to human IL-33 and anti human IgG1 Fc antibody. The expression level of scFv-Fc in plasmid PMH3(EN) was higher than that in plasmid pcDNA3.1. After four rounds of screening, the stably and high-level expressed cell strains were obtained. The expression level of the scFv-Fc was about 10 mg/L. The competitive ELISA results showed that the expressed scFv-Fc fusion proteins could inhibit the binding of IL-33 to ST2. Conclusion The anti-IL-33 scFv-Fc proteins were highly expressed in CHO k1 cells. PMID:27126936

  19. The novel anti-CD19 chimeric antigen receptors with humanized scFv (single-chain variable fragment) trigger leukemia cell killing.

    Science.gov (United States)

    Qian, Liren; Li, Dan; Ma, Lie; He, Ting; Qi, Feifei; Shen, Jianliang; Lu, Xin-An

    2016-01-01

    The molecular design of CARs (Chimeric Antigen Receptors), especially the scFv, has been a major part to use of CAR-T cells for targeted adoptive immunotherapy. To address this issue, we chose a vector backbone encoding a second-generation CAR based on efficacy of a murine scFv-based CAR. Next, we generated a panel of humanized scFvs and tested in vitro for their ability to direct CAR-T cells to specifically lyse, proliferate, and secrete cytokines in response to antigen-bearing targets. Furthermore, in a xenograft model of lymphoma, human T cells expressing humanized scFvs exhibited the same anti-tumor efficacy as those expressing murine scFv and prolonged survival compared with cells expressing control CAR. Therefore, we uncovered CARs expressing humanized scFv domain that contribute the similar enhanced antileukemic efficacy and survival in tumor bearing mice. These results provide the basis for the future clinical studies of CAR-T cells transduced with humanized scFv directed to CD19. PMID:26996927

  20. The inhibition of lung cancer cell growth by intracellular immunization with LC-1 ScFv

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    A monoclonal antibody, LC-l, recognizing lung cancer associated common antigens was obtained in authors' laboratory. Its single chain Fv fragment (ScFv) named LC-1 ScFv was constructed based on recombinant phage displayed techniques. For expression on cell membrane, LC-1 ScFv was cloned into pDisplay vector, which directed the cloned gene to express as cell membrane bound protein. The resulting plasmid was sequenced and then introduced by the lipofectin method into a lung adenocarcinoma cell line SPC-A-1. G418 resistant cells were obtained by G418 selection. After transfection, LC-1 ScFv expression was observed by Western blot analysis and the expression of cognate antigens was down-regulated as shown in ELISA assay. SPC-A-1-pDisplay-ScFv cells grew in vitro at lower speed than the control intact cells and the cells transfected with vacant vector. Flow cytometry analysis detected a substantial increase in G1 phase and decrease in S phase in population of SPC-A-1-pDisplay-ScFv cells compared to SPC-A-1 and SPC-A1-pDisplay cells. Semi-quantitative RT-PCR analysis showed that c-myc expression was down-regulated in SPC-A-1-pDisplay-ScFv cells. It seems that the antigens recognized by LC-1 may be in some way involved in a growth stimulating pathway and the antibody blocking of the function of the antigens shut down the pathway and thus down-regulate the expression of c-myc and growth of the cells.

  1. Rapid Purification of a New Humanized Single-chain Fv Antibody/Human Interleukin-2 Fusion Protein Reactive against HER2 Receptor

    Institute of Scientific and Technical Information of China (English)

    Wei-Yun ZHANG; Tak-Chun YIP; Cheuk-Sang KWOK

    2004-01-01

    Human embryonic kidney 293 cells were transfected with plasmid pcDNA-H520C9scFv-rhIL2 containing a chimeric cDNA encoding the humanized 520C9 scFv/recombinant human IL-2 fusion protein (H520C9scFv-rhIL-2). The transfected cells in plateau growing phase were cultured in serum-free medium for three days. The supernatant was collected, concentrated and purified using an affinity column packed with CNBr-activated Sepharose 4B coupled with anti-rhIL-2 mouse monoclonal antibody. The purified fusion protein was analyzed by ELISA, SDS-PAGE and Western blot. The fusion protein showed only one band in both silver stained electrophoresis gel and Western blot developed by ECL chemiluminescence system.Its molecular weight was confirmed to be about 45 kD. This fusion protein possessed binding specificity against p 185 positive SKOV3 and B 16/neu cells, and it might stimulate IL-2-dependent CTLL-2 cell proliferation as well.

  2. Embending Sustainability Dynamics in the Lean Construction Supply Chain Management

    Directory of Open Access Journals (Sweden)

    Sertyesilisik Begum

    2016-07-01

    Full Text Available The world’s habitat is being deteriorated despite of the precautions taken. Construction industry is among the industries which highly effect the environment adversely not only through its outputs but also through the construction process and its inputs. The main focus in dealing with the reduction of its footprint has been on sustainable building certificates which mainly analyse the output of the construction activies. There is need to analyse the construction supply chain as a whole and to embed sustainability dynamics in construction supply chain management. Lean construction project management contributes to the reduction of the environmental footprint of the construction industry, enabling reduction in waste, and increasing value added activities. For this reason, based on an in depth literature review, this paper analyses and establishes the principles of the integration of the sustainability dynamics into lean construction supply chain management.

  3. Selection, affinity maturation, and characterization of a human scFv antibody against CEA protein

    International Nuclear Information System (INIS)

    CEA is a tumor-associated antigen abundantly expressed on several cancer types, including those naturally refractory to chemotherapy. The selection and characterization of human anti-CEA single-chain antibody fragments (scFv) is a first step toward the construction of new anticancer monoclonal antibodies designed for optimal blood clearance and tumor penetration. The human MA39 scFv, selected for its ability to recognize a CEA epitope expressed on human colon carcinomas, was first isolated from a large semi-synthetic ETH-2 antibody phage library, panned on human purified CEA protein. Subsequently, by in vitro mutagenesis of a gene encoding for the scFv MA39, a new library was established, and new scFv antibodies with improved affinity towards the CEA cognate epitope were selected and characterized. The scFv MA39 antibody was affinity-maturated by in vitro mutagenesis and the new scFv clone, E8, was isolated, typed for CEA family member recognition and its CEACAM1, 3 and 5 shared epitope characterized for expression in a large panel of human normal and tumor tissues and cells. The binding affinity of the scFv E8 is in a range for efficient, in vivo, antigen capture in tumor cells expressing a shared epitope of the CEACAM1, 3 and 5 proteins. This new immunoreagent meets all criteria for a potential anticancer compound: it is human, hence poorly or not at all immunogenic, and it binds selectively and with good affinity to the CEA epitope expressed by metastatic melanoma and colon and lung carcinomas. Furthermore, its small molecular size should provide for efficient tissue penetration, yet give rapid plasma clearance

  4. Soluble Expression and Characterization of a New scFv Directed to Human CD123.

    Science.gov (United States)

    Moradi-Kalbolandi, Shima; Davani, Dariush; Golkar, Majid; Habibi-Anbouhi, Mahdi; Abolhassani, Mohsen; Shokrgozar, Mohammad Ali

    2016-04-01

    Leukemic cancer stem cells (LSCs), as a unique cell population in acute myeloid leukemia (AML) marked by CD123 overexpression, are thought to play a key role in relapsed AML after chemotherapy. Thus, CD123 is considered as a particularly important target candidate for antibody-derived diagnosis and therapy. In the present work, we constructed an immunized murine antibody phage display library and isolated the functional anti-CD123 Single-chain fragment variable (scFv) clones. We also introduced fusing variable light (VL) and heavy (VH) chains with a new 18-amino acid residue linker as an alternative to conventional linkers. CD123-specific phage clones were progressively enriched through 4 rounds of biopanning, validated by phage ELISA, and anti-CD123 scFv clones with highest affinity were produced in Escherichia coli. The expression and purification of soluble scFv were verified by Western blot, and the results were indicative of the functionality of our proposed linker. The purified scFv specifically recognized CD123 by ELISA and flow cytometry, without any cross-reactivity with other related cell markers. Affinity of anti-CD123 scFv was measured to be 6.9 × 10(-7) M, using the competitive ELISA. Our work, therefore, provides a framework for future studies involving biological functions and applications of our anti-CD123 scFv. It also reveals the feasibility of high throughput methods to isolate biomarker-specific scFvs. PMID:26749295

  5. Generation and Characterization of C305, a Murine Neutralizing scFv Antibody That Can Inhibit BLyS Binding to Its Receptor BCMA

    Institute of Scientific and Technical Information of China (English)

    Mei-Yun LIU; Wei HAN; Yan-Li DING; Tian-Hong ZHOU; Rui-Yang TIAN; Sheng-Li YANG; Hui LIU; Yi GONG

    2005-01-01

    B-lymphocyte stimulator (BLyS) is a member of the tumor necrosis factor (TNF) family and a key regulator of B cell response. Neutralizing single-chain fragment variable (scFv) antibody against BLyS binding to its receptor BCMA has the potential to play a prominent role in autoimmune disease therapy. A phage display scFv library constructed on pIII protein of M13 filamentous phage was screened using BLyS.After five rounds of panning, their binding activity was characterized by phage-ELISA. Nucleotide sequencing revealed that at least two different scFv gene fragments (C305 and D416) were obtained. The two different scFv gene fragments were expressed to obtain the soluble scFv antibodies, then the soluble scFv antibodies were characterized by means of competitive ELISA and in vitro neutralization assay. The results indicated that C305 is the neutralizing scFv antibody that can inhibit BLyS binding to its receptor BCMA.

  6. Construction of human single-chain variable fragment antibodies of medullary thyroid carcinoma and single photon emission computed tomography/computed tomography imaging in tumor-bearing nude mice.

    Science.gov (United States)

    Liu, Qiong; Pang, Hua; Hu, Xiaoli; Li, Wenbo; Xi, Jimei; Xu, Lu; Zhou, Jing

    2016-01-01

    Medullary thyroid carcinoma (MTC) is a rare tumor of the endocrine system with poor prognosis as it exhibits high resistance against conventional therapy. Recent studies have shown that monoclonal antibodies labeled with radionuclide have become important agents for diagnosing tumors. To elucidate whether single-chain fragment of variable (scFv) antibody labeled with 131I isotope is a potential imaging agent for diagnosing MTC. A human scFv antibody library of MTC using phage display technique was constructed with a capacity of 3x10(5). The library was panned with thyroid epithelial cell lines and MTC cell lines (TT). Western blotting and enzyme-linked immunosorbent assay (ELISA) were used to identify the biological characteristics of the panned scFv. Methyl thiazolyl tetrazolium (MTT) assay was also used to explore the optimal concentration of the TT cell proliferation inhibition rate. They were categorized into TT, SW480 and control groups using phosphate-buffered saline. Western blotting showed that molecular weight of scFv was 28 kDa, cell ELISA showed that the absorbance of TT cell group was significantly increased (P=0.000??) vs. the other three groups, and MTT assay showed that the inhibition rate between the two cell lines was statistically significantly different (P<0.05) when the concentration of scFv was 0.1, 1 and 10 µmol/l. The tumor uptake of 131I-scFv was visible at 12 h and clear image was obtained at 48 h using the single photon emission computed tomography. scFv rapidly and specifically target MTC cells, suggesting the potential of this antibody as an imaging agent for diagnosing MTC. PMID:26498224

  7. Construct validation of supply chain management in cooperative

    OpenAIRE

    Idris, Nurjihan; Arshad, Fatimah Mohamed; Radam, Alias; Ali, Noor Azman

    2009-01-01

    This study attempts to analyze construct in supply chain and to determine which construct contribute to performance of agricultural cooperatives in Malaysia. The primary data is collected via questionnaire from top level management of agricultural cooperatives using 5-item Likert scale. Factor analysis and structural equations modeling were used to analyze the data. Findings show that cooperatives places importance on quality and technology, logistic, supplier and governance. As a whole, supp...

  8. The molecular spin filter constructed from 1D organic chain

    International Nuclear Information System (INIS)

    We proposed a molecular spin filter, which is constructed from the 1D metallic organic chain (Fen+1(C6H4)n). The spin-polarized transport properties of the molecular spin filter are explored by combining density functional theory with nonequilibrium Green's function formalism. Theoretical results reveal that Fen+1(C6H4)n molecular chain exhibits robust spin filtering effect, and only the spin-down electrons can transmit through the molecular chain. At the given bias voltage window [−1 eV,1 eV], the calculated spin filter efficiency is close to 100% in the case of n≥3. We find that the effect of spin polarization origin from both Fen+1 and (C6H4)n. In addition, negative difference resistance behavior appears in Fen+1(C6H4)n molecular chain. The results can help us understand the spin transport properties of organic molecular chain. - Highlights: • Theoretical results reveal that Fen+1(C6H4)n molecular chain exhibits robust spin filtering effect. • The effect of spin polarization origin from both of Fen+1 and (C6H4)n. • Negative difference resistance behavior appears in Fen+1(C6H4)n molecular chain

  9. Transfection of an expressive construct including IgG1 and Fv1 genes in ovary cell line for infliximab expression

    Directory of Open Access Journals (Sweden)

    Zohreh Sarabinejad

    2016-04-01

    Full Text Available Background: Infeliximab is a form of chimeric antibody which neutralizes the most important inflammatory cytokine, TNF-a, in inflammatory disorders. The aim of current study was to pilot expression of chimeric infliximab in Chinese Hamster ovary (CHO cells. Methods: In this research study, pVITRO2-neo-mcs vector that consist of infliximab light chain and heavy chain was used to transform into the E.coli by CaCl2 method. The plasmid was then purified and transfected to cultured CHO cells by Lipofectamine 2000® (Invitrogen GmbH, Germany. Transfected cells were selected upon G-418 treatment after 2 weeks and the level of expression, based on standard curve, was measured using IgG ELISA kit after 48 hours for each clone. High level expressed clone was then cultured in roller bottles and recombinant chimeric product was purified by protein A affinity chromatography. The purity of the product was analyzed by 10% gel SDS-PAGE from eluted samples. The efficacy of the purification was analyzed by ELISA before and after purification step. This article is a master's student thesis from February 2015 to August 2016 in pharmaceutical technology development center, Tehran University of Medical Sciences, Tehran, Iran. Results: The purified plasmid was analyzed on 2% agarose gel. After selective pressure of G-418, 10 stable transfect clones were assessed for infliximab secretion by IgG ELISA kit at 450 nm. The maximum and minimum expression which detected by ELISA were 23 ng/ml and 6 ng/ml, respectively. The band width of infliximab fraction during purification procedure was observed at 0.7-0.8 min. The efficiency of the purification by ELISA was 70%. On SDS-PAGE analysis, two bands, 25 and 50 kDa, respect to light and heavy chains of Infliximab, was confirmed the expression of recombinant protein. Conclusion: In the current study, the construct for infliximab monoclonal antibody production was designed using genetic engineering techniques and the expression

  10. Constructing 1/ωα noise from reversible Markov chains

    Science.gov (United States)

    Erland, Sveinung; Greenwood, Priscilla E.

    2007-09-01

    This paper gives sufficient conditions for the output of 1/ωα noise from reversible Markov chains on finite state spaces. We construct several examples exhibiting this behavior in a specified range of frequencies. We apply simple representations of the covariance function and the spectral density in terms of the eigendecomposition of the probability transition matrix. The results extend to hidden Markov chains. We generalize the results for aggregations of AR1-processes of C. W. J. Granger [J. Econometrics 14, 227 (1980)]. Given the eigenvalue function, there is a variety of ways to assign values to the states such that the 1/ωα condition is satisfied. We show that a random walk on a certain state space is complementary to the point process model of 1/ω noise of B. Kaulakys and T. Meskauskas [Phys. Rev. E 58, 7013 (1998)]. Passing to a continuous state space, we construct 1/ωα noise which also has a long memory.

  11. Last Planner and Critical Chain in Construction Management: Comparative Analysis

    OpenAIRE

    Koskela, Lauri; Stratton, Roy; Koskenvesa, Anssi

    2010-01-01

    This paper endeavours to compare the Last Planner System of production control and the Critical Chain production management method. This comparison is carried out in the context of construction management. The original prescription and the evolution of the practice are examined regarding both approaches, and the similarities and differences are noted. Based on these considerations, gaps in the two approaches are identified and the potential of a synthesis of them is explored.

  12. Recombinant norovirus-specific scFv inhibit virus-like particle binding to cellular ligands

    Directory of Open Access Journals (Sweden)

    Hardy Michele E

    2008-01-01

    Full Text Available Abstract Background Noroviruses cause epidemic outbreaks of gastrointestinal illness in all age-groups. The rapid onset and ease of person-to-person transmission suggest that inhibitors of the initial steps of virus binding to susceptible cells have value in limiting spread and outbreak persistence. We previously generated a monoclonal antibody (mAb 54.6 that blocks binding of recombinant norovirus-like particles (VLP to Caco-2 intestinal cells and inhibits VLP-mediated hemagglutination. In this study, we engineered the antigen binding domains of mAb 54.6 into a single chain variable fragment (scFv and tested whether these scFv could function as cell binding inhibitors, similar to the parent mAb. Results The scFv54.6 construct was engineered to encode the light (VL and heavy (VH variable domains of mAb 54.6 separated by a flexible peptide linker, and this recombinant protein was expressed in Pichia pastoris. Purified scFv54.6 recognized native VLPs by immunoblot, inhibited VLP-mediated hemagglutination, and blocked VLP binding to H carbohydrate antigen expressed on the surface of a CHO cell line stably transfected to express α 1,2-fucosyltransferase. Conclusion scFv54.6 retained the functional properties of the parent mAb with respect to inhibiting norovirus particle interactions with cells. With further engineering into a form deliverable to the gut mucosa, norovirus neutralizing antibodies represent a prophylactic strategy that would be valuable in outbreak settings.

  13. Porter's value chain (construction, deconstruction, reconstruction and values management

    Directory of Open Access Journals (Sweden)

    E.V. Krykavskyy

    2015-06-01

    Full Text Available The aim of the article. The phases of the Porter's value chain are distinguished: construction of chain value – Porters model (Stage 1; deconstruction – identifying contradictions, disorganizing elements of unnecessary processes that do not add value (Stage 2; reconstruction (synthesis – creates a new value chain (Stage 3. The results of the analysis. The principles of convergence of value and supply chains are identified and the need to focus on supply chain performance is proved. The dependence of design value chain/supply chain on the chosen general strategy and the need to subject its reconstruction process of deconstruction + reconstruction in case of changes or modifications as to this strategy is grounded. The impacts on the creation of value in the Porter’s chain are accepted. The need to identify for each working block of micro factors that affect the financial factors (macro factors that is net and operating profit, investments into fixed and working capital, the cost of companies capital is proved. Such classification allows to determine what types of management actions or motivations for managers of lower level must be selected. It is revealed that in addition to measurable factors-generators on cost to the value chain the indirect (mediated factors have an impact, especially when the management is replaced by objectives and values of enterprise management use. It is proved that the problem of environment and society in the concept of sustainable development, social responsibility, marketing value of 3.0 outlines another group of factors influencing the Porter’s value chain, as evidenced by the current trend of prioritization of human values. It is proved that companies, especially global ones, give the increasing domination of human values in B2B and B2C transactions at the maturity stage in their life cycle in the structure of forced strategic objectives and make adjustments by modifying the cited above vision and mission

  14. Surface display of an anti-DEC-205 single chain Fv fragment in Lactobacillus plantarum increases internalization and plasmid transfer to dendritic cells in vitro and in vivo

    OpenAIRE

    Christophe, Michon; Kuczkowska, Katarzyna; Langella, Philippe; Eijsink, Vincent; Mathiesen, Geir; Chatel, Jean-Marc

    2015-01-01

    Background Lactic acid bacteria (LAB) are promising vehicles for delivery of a variety of medicinal compounds, including antigens and cytokines. It has also been established that LAB are able to deliver cDNA to host cells. To increase the efficiency of LAB-driven DNA delivery we have constructed Lactobacillus plantarum strains targeting DEC-205, which is a receptor located at the surface of dendritic cells (DCs). The purpose was to increase uptake of bacterial cells, which could lead to impro...

  15. Targeting immune effector molecules to human tumor cells through genetic delivery of 5T4-specific scFv fusion proteins.

    Science.gov (United States)

    Myers, Kevin A; Ryan, Matthew G; Stern, Peter L; Shaw, David M; Embleton, M Jim; Kingsman, Susan M; Carroll, Miles W

    2002-11-01

    Although several clinical trials have shown beneficial effects by targeting tumor-associated antigens (TAAs) with monoclonal antibodies, a number of issues, including poor penetration of the tumor mass and human antimouse antibody responses, remain. The use of recombinant single-chain Fv (scFv) fragments has the potential to address these and other issues while allowing the addition of different effector functions. To develop therapeutic strategies that recruit both humoral and cellular arms of the immune response, we have constructed chimeric proteins linking either the human IgG1 Fc domain or the extracellular domain of murine B7.1 to a scFv specific for the oncofetal glycoprotein, 5T4. This TAA is expressed by a wide variety of carcinomas and is associated with metastasis and poorer clinical outcome. We have engineered retroviral constructs that produce fusion proteins able to interact simultaneously with both 5T4-positive cells and with the receptor/ligands of the immune effector moieties. Genetic delivery through a murine leukemia virus vector to 5T4-positive tumor cells results in the secreted scFv fusion protein binding to the cell surface. Furthermore, the scFv-HIgG1 fusion protein is able to direct lysis of 5T4-expressing human tumor cell lines through antibody-dependent cell cytotoxicity, indicating its potential as a gene therapy for human cancers. PMID:12386827

  16. How to construct the spin chains with perfect state transfer

    CERN Document Server

    Vinet, Luc

    2011-01-01

    It is shown how to systematically construct the $XX$ quantum spin chains with nearest-neighbor interaction that allow perfect state transfer (PST). Sets of orthogonal polynomials (OPs) are in correspondence with such systems. The key observation is that for any admissible one-excitation energy spectrum, the weight function of the associated OPs is uniquely prescribed. This entails the complete characterization of these PST models with the mirror symmetry property arising as a corollary. A simple and efficient algorithm to obtain the corresponding Hamiltonians is presented. A new model connected to a special case of the symmetric $q$-Racah polynomials is offered. It is also explained how additional models with PST can be derived from from a parent system by removing energy levels from the one-excitation spectrum of the latter. This is achieved through Christoffel transformations and is also completely constructive in regards to the Hamiltonians.

  17. Isolation of a high affinity scFv from a monoclonal antibody recognising the oncofoetal antigen 5T4.

    Science.gov (United States)

    Shaw, D M; Embleton, M J; Westwater, C; Ryan, M G; Myers, K A; Kingsman, S M; Carroll, M W; Stern, P L

    2000-12-15

    The oncofoetal antigen 5T4 is a 72 kDa glycoprotein expressed at the cell surface. It is defined by a monoclonal antibody, mAb5T4, that recognises a conformational extracellular epitope in the molecule. Overexpression of 5T4 antigen by tumours of several types has been linked with disease progression and poor clinical outcome. Its restricted expression in non-malignant tissue makes 5T4 antigen a suitable target for the development of antibody directed therapies. The use of murine monoclonal antibodies for targeted therapy allows the tumour specific delivery of therapeutic agents. However, their use has several drawbacks, including a strong human anti-mouse immune (HAMA) response and limited tumour penetration due to the size of the molecules. The use of antibody fragments leads to improved targeting, pharmacokinetics and a reduced HAMA. A single chain antibody (scFv) comprising the variable regions of the mAb5T4 heavy and light chains has been expressed in Escherichia coli. The addition of a eukaryotic leader sequence allowed production in mammalian cells. The two 5T4 single chain antibodies, scFv5T4WT19 and LscFv5T4, described the same pattern of 5T4 antigen expression as mAb5T4 in normal human placenta and by FACS. Construction of a 5T4 extracellular domain-IgGFc fusion protein and its expression in COS-7 cells allowed the relative affinities of the antibodies to be compared by ELISA and measured in real time using a biosensor based assay. MAb5T4 has a high affinity, K(D)=1.8x10(-11) M, as did both single chain antibodies, scFv5T4WT19 K(D)=2.3x10(-9) M and LscFv5T4 K(D)=7.9x10(-10) M. The small size of this 5T4 specific scFv should allow construction of fusion proteins with a range of biological response modifiers to be prepared whilst retaining the improved pharmacokinetic properties of scFvs. PMID:11113573

  18. Primary structure and functional scFv antibody expression of an antibody against the human protooncogen c-myc.

    Science.gov (United States)

    Fuchs, P; Breitling, F; Little, M; Dübel, S

    1997-06-01

    The immunoglobulin heavy- and light-chain variable region (Vh and Vl) genes were isolated from Myc1-9E10 hybridoma cells, which secreted monoclonal antibody against human oncogen c-myc. The expression vector pOPE52-c-myc was constructed for the recombinant production in E. coli. A 30 kDa single chain fragment (scFv) expression product was found in the periplasmic space by SDS-PAGE and immunoblotting. A significant fraction was processed correctly as demonstrated with an antiserum recognizing the processed aminoterminus only. The specific binding of the scFv fragment to the peptide epitope of the maternal monoclonal antibody was demonstrated and the primary sequence of the variable regions was determined. Sequence comparison with previously published partial Vh and Vl sequences from this hybridoma cell line revealed a genetic heterogeneity for the light chain variable region. The potential use of this scFv as a new tool for detection and purification of tagged proteins, for adding costimulatory signals to the surface of cancer cells as well as for analyzing c-myc function in the living cell by cytoplasmic expression is discussed. PMID:9219032

  19. An efficient method for variable region assembly in the construction of scFv phage display libraries using independent strand amplification

    OpenAIRE

    Sotelo, Pablo H.; Collazo, Noberto; Zuñiga, Roberto; Gutiérrez-González, Matías; Catalán, Diego; Ribeiro, Carolina Hager; Aguillón, Juan Carlos; MOLINA, MARÍA CARMEN

    2012-01-01

    Phage display library technology is a common method to produce human antibodies. In this technique, the immunoglobulin variable regions are displayed in a bacteriophage in a way that each filamentous virus displays the product of a single antibody gene on its surface. From the collection of different phages, it is possible to isolate the virus that recognizes specific targets. The most common form in which to display antibody variable regions in the phage is the single chain variable fragment...

  20. Demand uncertainty in construction supply chains: a discrete event simulation study

    OpenAIRE

    C Vidalakis; J.E. Tookey; Sommerville, J

    2013-01-01

    The delivery of construction projects is typically an assembly operation involving a high number of subassemblies and materials brought on site by the supply chain. However, although supply chain management in construction has attracted significant attention, paradoxically little focus has been placed on construction supply networks and operations. This paper places emphasis on supply chain operations by looking at the logistics function of construction material suppliers. Specifically, the p...

  1. Construction of human Alzheimer's disease specific single chain antibodies library%人源性阿尔茨海默病噬菌体单链抗体库的构建

    Institute of Scientific and Technical Information of China (English)

    李楠; 王建平

    2015-01-01

    Objective To construct the human Alzheimer's disease (AD) specific single chain anti‐bodies (scFv) library for sceening human AD scFv to Aβ1‐42 oligomers .Methods RNA was isola‐ted from 40 ml peripheral blood taken from 18 AD patients .Variable heavy (V H ) and variable light (VL ) genes were amplified by RT‐PCR and linked to the scFv fragments that were then cloned into the phage vector pCANTAB5E .The scFv library was constructed by electroporating E .coli TG1 cells into the pCANTAB5E and rescuing the assistant phage M13K07 .Results The total RNA was extracted .The VH and VL genes were amplified .Electropharesis showed that the length of VH and VL genes was 360bp and 300bp respectively and the length of linked scFv was 750bp .The 2 .4 × 109 scFv library was thus constructed and identified by BstN I digestion .Elec‐tropharesis showed that the length of BstN I‐digested scFv fragments varied ,indicating that the library is of a good diversity .Conclusion The human AD phage scFv library we constructed lays a foundation for screening the antibodies to Aβ1‐42 oligomers and the treatment of AD .%目的:构建人源性阿尔茨海默病(AD)噬菌体单链抗体(scFv)库,为筛选β淀粉样蛋白(Aβ1‐42)的人源性特异性抗体奠定基础。方法采集18例AD患者的外周血40 ml ,提取总RNA ,应用RT‐PCR法得到人抗体可变区重链(V H )和可变区轻链(V L )基因。将 V H 和 V L 由连接肽连接得到 scFv 片段,将所得片段双酶切后,克隆至pCANTAB5E噬菌体载体,大肠埃希菌TG1感受态细胞经电击转化,辅助噬菌体 M13K07拯救后构建scFv型噬菌体抗体库。结果总RNA经逆转录PCR扩增VH和VL可变区基因的凝胶电泳显示,PCR产物长度分别为360 bp和300 bp ,其连接形成的scFv片段长度为750 bp ,最终构建了库容为2.4×109的scFv库。BstNⅠ酶切鉴定构建的scFv库,经凝胶电泳可见,酶切片段长度差异性大,

  2. Isolation of ScFv antibodies of rP27Kip1 from phage display libraries constructed from immunized and non-immunized repertoires

    Institute of Scientific and Technical Information of China (English)

    曹跃琼; 乔守怡; 袁有忠; 黄建生; 赵寿元

    1999-01-01

    Through mRNA extract, RT and a series of PCR, phage antibody libraries were constructed from rP27Kiplimmunized and non-immunized mice. After only one round of selection with rP27Kipl, clones from each library were chosen randomly and digested by Taq I and Hinf I. 11 of 64 clones from the immunized animal had consistent restriction pattern, while none of the 64 clones from the non-immunized animal had, except that one had the same fragments pattern as that of the 11 clones. The 12 fragments were expressed in E. coli BL21(DE3)/pET-28b(+) system. ELISA showed that some of the fragments could bind to rP27Kipl specifically. All these results implied that specific antibody can be obtained by genetic engineering without hybridoma or many rounds of growth and panning selection.

  3. Gigantoxin-4-4D5 scFv is a novel recombinant immunotoxin with specific toxicity against HER2/neu-positive ovarian carcinoma cells.

    Science.gov (United States)

    Lv, Xinxin; Zhang, Jian; Xu, Rui; Dong, Yuguo; Sun, Aiyou; Shen, Yaling; Wei, Dongzhi

    2016-07-01

    Immunotoxins are a new class of antibody-targeted therapy in clinical development. Traditional immunotoxins that are constructed from the toxins of plants or bacteria need to be internalized to the cytoplasm and thus have limited antitumor efficacy. In the present study, we combined a recently reported sea anemone cytolysin Gigantoxin-4 with an anti-HER2/neu single-chain variable fragment 4D5 scFv to construct a novel immunotoxin. We fused a SUMO tag to the N-terminus of Gigantoxin-4-4D5 scFv and it was successfully expressed in Escherichia coli strain BL21 (DE3) in a soluble form. After purification, the purity of Gigantoxin-4-4D5 scFv reached 96 % and the yield was 14.3 mg/L. Our results demonstrated that Gigantoxin-4-4D5 scFv exerted a highly cytotoxic effect on the HER2/neu-positive ovarian carcinoma SK-OV-3 cell line. And the hemolytic activity was weaker, making it safe for normal cells. The results of immunofluorescence analysis showed that this novel immunotoxin could specifically bind to SK-OV-3 cells with no recognition of human embryonic kidney 293 cells. Scanning electron microscope observations and extracellular lactate dehydrogenase activity indicated that it could induce necrosis in SK-OV-3 cells by disrupting the cell membrane. Moreover, it could also mediate apoptosis of SK-OV-3 cells. PMID:27063011

  4. Fuzzy Optimization of Construction Engineering Project Schedule Based on Critical Chain Management

    OpenAIRE

    Liu, Jianbing; Ren, Hong; Junshu DU

    2013-01-01

    Critical chain management was the very important theory innovation in the development of construction project schedule management field in recent years. Compared with the traditional methods of construction project schedule management, the methods of critical chain management considered resource constraints into construction project schedule management, the construction project cycle was shortened and the efficiency was enhanced by using the dynamic thinking and circulation pattern to manage ...

  5. Construction of Network Management Information System of Agricultural Products Supply Chain Based on 3PLs

    OpenAIRE

    Gao, Shujin; Yang, Qiangxian

    2010-01-01

    The necessity to construct the network management information system of 3PLs agricultural supply chain is analyzed, showing that 3PLs can improve the overall competitive advantage of agricultural supply chain. 3PLs changes the homogeneity management into specialized management of logistics service and achieves the alliance of the subjects at different nodes of agricultural products supply chain. Network management information system structure of agricultural products supply chain based on 3PL...

  6. Measuring Safety Attitude Differences in the Construction Supply Chain

    OpenAIRE

    Saunders, Lance Walter

    2013-01-01

    Construction worker safety is normally a construction activity in the United States, even though there is an emerging body of literature discussing the positive effects of considering safety earlier in the construction lifecycle.  This literature has discussed the fragmentation in terms of safety attitudes between owners and designers and those carrying out the construction of a project.  Quantitatively identifying the specific areas that the differences exist in terms of safety attitudes bet...

  7. Production and characterization of a recombinant anti-MUC1 scFv reactive with human carcinomas.

    OpenAIRE

    Denton, G; Sekowski, M.; Spencer, D. I.; Hughes, O. D.; Murray, A; Denley, H; Tendler, S. J.; Price, M. R.

    1997-01-01

    Recombinant single-chain fragments (scFv) of the murine anti-MUC1 monoclonal antibody C595 have been produced using the original hybridoma cells as a source of variable heavy (V(H))- and variable light (V(L))-chain-encoding antibody genes. The use of the polymerase chain reaction (PCR), bacteriophage (phage) display technology and gene expression systems in E. coli has led to the production of soluble C595 scFv. The scFv has been purified from the bacterial supernatant by peptide epitope affi...

  8. The Rural Information-based Construction under the Perspective of Expanding Agricultural Industrial Chain

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    On the basis of expounding connotation and significance of expansion of agricultural industrial chain, coupled with the connotation of rural informatization connotation, this article analyses the role of rural informatization in expanding agricultural industrial chain: it can enhance market competitiveness of industry chain, improve the operational efficiency of industry chain, and promote the income and quality of farmers in industry chain. Under the perspective of expanding agricultural industrial chain, this article puts forwards thinking about the construction of rural informatization as follows: first, give full play to the leading role of the government; second, strengthen the construction of information-based network facility; third, integrate information resources in rural areas, and improve the quality of information; fourth, build comprehensive information service platform in rural areas; fifth, improve organizational level of production and management of individual farmers; sixth, strengthen the construction of information-based personnel in rural areas; seventh, strengthen publicity and training, promote overall cultural quality and information awareness of farmers.

  9. 抗阿尔茨海默病Aβ人源单链抗体基因的筛选和表达%Cloning and expression of human single-chain Fv antibody against Aβ1-42 peptide involved in Alzheimer disease

    Institute of Scientific and Technical Information of China (English)

    贾静; 刘喆; 赵雪梅; 梁平

    2008-01-01

    目的 从人源噬菌体单链抗体库中筛选与阿尔茨海默病发病中起关键作用的β淀粉样多肽(Aβ)1-42特异性结合的单链可变区抗体(scFv)基因,利用原核生物大肠杆菌进行可溶性表达,以获得抗AB1-42人源抗体.方法 利用噬菌体展示技术对人源噬菌体单链抗体库进行多轮富集,以Aβ1-42为抗原经酶联免疫吸附(ELISA)检测,筛选与Aβ1-42特异性结合的阳性噬菌体克隆,再用阳性噬菌体感染E.coli HB2151进行可溶性表达,经SDS-PAGE、Western blot及ELISA法检测scFv单抗的可溶性表达水平及抗原结合活性.并对阳性scFv抗体基因进行基因测序鉴定.结果 获得了7个特异性的抗Aβ1-42 scFv阳性噬菌体克隆,其中4个克隆ELISA检测吸光度值(A值)高于阴性对照5倍以上;其中1株(A 10)获得可溶性单链抗体的成功表达,表达产物主要位于菌体中,ELISA效价显示在全菌蛋白中A值高于对照5倍以上.其相对分子质量约为33 000.DNA测序表明所获抗体的可变区基因属于scFv抗体基因,推导得到的氨基酸序列具有典型的抗体可变区结构.结论 用人源噬菌体单链抗体库筛选出抗Aβ1-42的人源抗体单链可变区基因,并成功表达了具有抗原结合活性的可溶性单链抗体,为进一步研究阿尔茨海默病的发病机制和治疗奠定了基础.%Objective Screening of antibody clones specific for β amyloid peptide 1-42 from human phage-display single-chain Fv(scFv)antibody library,and to clone the antibody gene and to express it in a bacterial system,with an ultimate intention to obtain human anti-Aβ1-42 antibody for Alzheimer disease(AD) therapy.Methods β amyloid peptide 1-42 was bound on the solid surface of 96 wells plate as the antigen for the binding antibody clones from a human phage-display scFv antibody library.After four rounds of biopanning,random,well-separated colonies were identified by ELISA test.The specific positive phage clones were

  10. Construction and Application of a Refined Hospital Management Chain.

    Science.gov (United States)

    Lihua, Yi

    2016-01-01

    Large scale development was quite common in the later period of hospital industrialization in China. Today, Chinese hospital management faces such problems as service inefficiency, high human resources cost, and low rate of capital use. This study analyzes the refined management chain of Wuxi No.2 People's Hospital. This consists of six gears namely, "organizational structure, clinical practice, outpatient service, medical technology, and nursing care and logistics." The gears are based on "flat management system targets, chief of medical staff, centralized outpatient service, intensified medical examinations, vertical nursing management and socialized logistics." The core concepts of refined hospital management are optimizing flow process, reducing waste, improving efficiency, saving costs, and taking good care of patients as most important. Keywords: Hospital, Refined, Management chain PMID:27180468

  11. Supply chain for new construction: buy where we build

    International Nuclear Information System (INIS)

    A global supply strategy has been placed by Westinghouse in order to face the current and future constructions. And in this way, Westinghouse will be able to take advantage of benefit of Local Supply. this article shows the Westinghouse Global Supply Strategy, Local Supply considerations, the current state of supplier calcifications, and finally Memorandums of Understanding and Alliances agreed with local suppliers around the world to provide a suitable solution for the new construction needs. (Author)

  12. An Adaptively Constructed Algebraic Multigrid Preconditioner for Irreducible Markov Chains

    OpenAIRE

    Brannick, James; Kahl, Karsten; Sokolovic, Sonja

    2014-01-01

    The computation of stationary distributions of Markov chains is an important task in the simulation of stochastic models. The linear systems arising in such applications involve non-symmetric M-matrices, making algebraic multigrid methods a natural choice for solving these systems. In this paper we investigate extensions and improvements of the bootstrap algebraic multigrid framework for solving these systems. This is achieved by reworking the bootstrap setup process to use singular vectors i...

  13. Fuzzy Optimization of Construction Engineering Project Schedule Based on Critical Chain Management

    Directory of Open Access Journals (Sweden)

    Jianbing LIU

    2013-07-01

    Full Text Available Critical chain management was the very important theory innovation in the development of construction project schedule management field in recent years. Compared with the traditional methods of construction project schedule management, the methods of critical chain management considered resource constraints into construction project schedule management, the construction project cycle was shortened and the efficiency was enhanced by using the dynamic thinking and circulation pattern to manage whole project. In view of the progress of critical chain management of the article, fuzzy theory  was used to calculate and optimize the buffer zone sizes with the determination of the buffer size of  the existing resources in certain critical chain management so as to shorten the cycle..

  14. Strategies for integrating design and construction and operations and maintenance supply chains in Singapore

    OpenAIRE

    Ling, FYY; Toh, BGY; Kumaraswamy, MM; Wong, KWK

    2014-01-01

    Purpose – The purpose of this paper is to investigates strategies for achieving better integration between the design and construction (DC) and operation and maintenance (OM) supply chains in Singapore. The specific objectives are to: discover the goals that stakeholders want to achieve in integrating the supply chains; identify the stakeholders that play important integration role in each supply chain; and investigate the effective strategies that may yield better integration of the supply ...

  15. Enzymic construction of maltosaccharide chains on a heart protein

    International Nuclear Information System (INIS)

    The authors have reported that when 100,000 g pellets of rabbit-heart and rabbit-muscle homogenates are incubated with UDP(14C)glucose, the sugar is incorporated into a protein with Mr 40 KDa. They suggested that these in vitro observations corresponded to the initial stage in the synthesis of glycogen on a protein that they have named glycogenin and which in rabbit muscle appears to be covalently linked to the glycogen via tyrosine residues. The following new observations support the role of a protein as the precursor of glycogen and suggest that glycogen-free glycogenin is present in heart tissue. (1) The (14C)glucose residues added to the heart protein can be removed with glycogenolytic enzymes that hydrolyse 1,4-alpha-glucosidic bonds and therefore constitute synthetic maltosaccharide chains. (2) The newly added glucose residues appear to be attached to pre-existing glucose residues on the protein. Chain elongation does not proceed beyond a few glucose residues. (3) The further relevance of these observations to glycogen synthesis shown by a Western blot in which the radioglucosylated heart protein was found to cross-react with polyclonal antibody to glycogenin obtained from rabbit-muscle glycogen

  16. Synergistic capture of Clostridium botulinum Type A neurotoxin by scFv antibodies to novel epitopes

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Sean A.; Barr, John R.; Kalb, Suzanne R.; Marks, James D.; Baird, Cheryl L.; Cangelosi, Gerard A.; Miller, Keith D.; Feldhaus, Michael J.

    2011-10-01

    A non-immune library of human single chain fragment variable (scFv) antibodies displayed on Saccharomyces cerevisiae was screened for binding to the Clostridium botulinum neurotoxin serotype A binding domain [BoNT/A (Hc)] with the goal of identifying scFv to novel epitopes. To do this, an antibody-mediated labeling strategy was used in which antigen-binding yeast clones were selected after labeling with previously characterized monoclonal antibodies (MAbs) specific to the Hc. Twenty unique scFv clones were isolated that bound Hc. Of these, three also bound to full-length BoNT/A toxin complex with affinities ranging from 5 nM to 170 nM. Epitope binning showed that the three unique clones recognized at least two epitopes that were distinct from one another and from the detection MAbs. After production in E. coli, the scFv were coupled to magnetic particles and tested for their ability to capture BoNT/A holotoxin using an Endopep-MS assay. In this assay, toxin captured by scFv coated magnetic particles was detected by incubation of the complex with a peptide containing a BoNT/A-specific cleavage sequence. Mass spectrometry was used to detect the ratio of intact peptide to cleavage products as evidence for toxin capture. When tested individually, each of the scFv showed a weak positive Endopep-MS result. However, when the particles were coated with all three scFv simultaneously, they exhibited significantly higher Endopep-MS activity, consistent with synergistic binding. These results demonstrate novel approaches toward the isolation and characterization of scFv antibodies specific to unlabeled antigen. They also provide evidence that distinct scFv antibodies can work synergistically to increase the efficiency of antigen capture onto a solid support.

  17. Constructing Chains of Enablers for Alternative Economic Futures

    DEFF Research Database (Denmark)

    Hull Kristensen, Peer

    2016-01-01

    that have made possible novel forms of work-organization and business models. These institutional enablers are capacitating on the "supply side" by enabling labor to take active part in shaping enterprises supported by social welfare services (training, child- and eldercare, support for housing, etc...... economies. This article illustrates a way of researching alternative economic futures by identifying chains of enablers in Denmark and other Nordic countries by which society and business can co-develop and capture capabilities to take on new roles in globalization. Focus is on institutional enablers.......). Being generally inclusive of social movements, welfare states has also helped identify new needs on the "demand side" such as child- and eldercare, environmental protection, alternative energy and energy-saving, health, and city planning. This is illustrated by a number of firms that supply products...

  18. On Construction of Supply Chain System for China’s Modern Agricultural Products

    Institute of Scientific and Technical Information of China (English)

    Wanjiang; WANG

    2015-01-01

    In view of drawbacks in supply chain of China’s traditional agricultural products,this paper proposed building supply chain system for modern agricultural products: taking informationization as basis,channel system as core,organization system as support,and service system and safety system as guarantee,to promote high efficient operation of the supply chain system. The channel system stresses alliance and integration of channel system,informationization of channel management,and terminalization of channel operation; the organization system stresses organization,large scale,group,and brand of participant entities; service system stresses construction of service means,service platform,and operation mechanism; safety system stresses building quality safety based agricultural product supply chain management mode. In order to ensure high efficient operation of supply chain for modern agricultural products,it is required to straighten out supply chain management system,actively cultivate core enterprises of supply chain,strengthen information construction of supply chain,select suitable supply chain mode,and improve benefit allocation mechanism.

  19. Network design and operational modelling for construction green supply chain management

    Directory of Open Access Journals (Sweden)

    Pengfei Zhou Dong Chen

    2013-01-01

    Full Text Available Based on studying organizational structure of Construction Green Supply Chain Management (CGSCM, a mathematical programming model of CGSCM was proposed. The model aimed to maximize the aggregate profits of normalized construction logistics, the reverse logistics and the environmental performance. Numerical experiments show that the proposed approach can improve the aggregate profit effectively. In addition, return ratio, subsidies from governmental organizations, and environmental performance were analyzed for CGSCM performance. Herein, the proper return, subsidy and control strategy could optimize construction green supply chain.

  20. Efficient silkworm expression of single-chain variable fragment antibody against ginsenoside Re using Bombyx mori nucleopolyhedrovirus bacmid DNA system and its application in enzyme-linked immunosorbent assay for quality control of total ginsenosides.

    Science.gov (United States)

    Sakamoto, Seiichi; Pongkitwitoon, Benyakan; Nakamura, Seiko; Maenaka, Katsumi; Tanaka, Hiroyuki; Morimoto, Satoshi

    2010-09-01

    A single-chain variable fragment (scFv) antibody against ginsenoside Re (G-Re) have been successfully expressed in the silkworm larvae using Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid DNA system. The baculovirus donor vector for expression of scFv against G-Re (GRe-scFv) was constructed to contain honeybee melittin signal sequence to accelerate secretion of the recombinant GRe-scFv into the haemolymph of silkworm larvae. Functional recombinant GRe-scFv was purified by cation exchange chromatography followed by immobilized metal ion affinity chromatography. The yield of purified GRe-scFv was 6.5 mg per 13 silkworm larvae, which is equivalent to 650 mg/l of the haemolymph, exhibiting extremely higher yield than that expressed in Escherichia coli (1.7 mg/l of culture medium). It was revealed from characterization that GRe-scFv retained similar characteristic of the parental monoclonal antibody (MAb) against G-Re (MAb-4G10), making it possible to develop indirect competitive enzyme-linked immunosorbent assay (icELISA) for quality control of total ginsenosides in various ginsengs. The detectable range for calibration of G-Re by developed icELISA shows 0.05-10 microg/ml. These results clearly suggested that the silkworm expression system is quite useful for the expression of functional scFv that frequently required time- and cost-consuming re-folding when it expressed in E. coli. PMID:20592135

  1. DIRECTIONS FOR FUTURE CONSTRUCTION SUPPLY CHAIN MANAGEMENT RESEARCH IN NEW ZEALAND: A REAL OPTIONS PERSPECTIVE

    Directory of Open Access Journals (Sweden)

    Tran, Van

    2012-04-01

    Full Text Available Real Options (RO has been a universally accepted concept in a number of major industries. However, its use in the construction supply chain management (CSCM sector has been limited. Some rare supply chain management RO studies have shown a number of limitations. First, there is a lack of a rigorous theoretical RO framework pertaining specifically to CSCM. All such supply chain management RO studies are based off RO theories or models developed for other sectors (engineering, infrastructure, natural resources. And second, attempts to extend real option to wider uses in CSCM seem premature at the present. This paper reviews all recent literature pertaining to real options and real options applied specifically to the construction supply chain management area. The study proposes a research programme pertaining to CSCM in New Zealand in order to enhance the current understanding of RO in this area and in the process develop a comprehensive theory for the RO application in New Zealand CSCM.

  2. Construction of Network Management Information System of Agricultural Products Supply Chain Based on 3PLs

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    The necessity to construct the network management information system of 3PLs agricultural supply chain is analyzed,showing that 3PLs can improve the overall competitive advantage of agricultural supply chain.3PLs changes the homogeneity management into specialized management of logistics service and achieves the alliance of the subjects at different nodes of agricultural products supply chain.Network management information system structure of agricultural products supply chain based on 3PLs is constructed,including the four layers (the network communication layer,the hardware and software environment layer,the database layer,and the application layer) and 7 function modules (centralized control,transportation process management,material and vehicle scheduling,customer relationship,storage management,customer inquiry,and financial management).Framework for the network management information system of agricultural products supply chain based on 3PLs is put forward.The management of 3PLs mainly includes purchasing management,supplier relationship management,planning management,customer relationship management,storage management and distribution management.Thus,a management system of internal and external integrated agricultural enterprises is obtained.The network management information system of agricultural products supply chain based on 3PLs has realized the effective sharing of enterprise information of agricultural products supply chain at different nodes,establishing a long-term partnership revolving around the 3PLs core enterprise,as well as a supply chain with stable relationship based on the supply chain network system,so as to improve the circulation efficiency of agricultural products,and to explore the sales market for agricultural products.

  3. Novel immunocytokine IL12-SS1 (Fv) inhibits mesothelioma tumor growth in nude mice.

    Science.gov (United States)

    Kim, Heungnam; Gao, Wei; Ho, Mitchell

    2013-01-01

    Mesothelin is a glycosylphosphatidylinositol-anchored glycoprotein that is highly expressed on the cell surface of malignant mesothelioma. Monoclonal antibodies against mesothelin are being evaluated for the treatment of mesothelioma. Immunocytokines represent a novel class of armed antibodies. To provide an alternative approach to current mesothelin-targeted antibody therapies, we have developed a novel immunocytokine based on interleukin-12 (IL12) and the SS1 Fv specific for mesothelin. IL12 possesses potent anti-tumor activity in a wide variety of solid tumors. The newly-developed recombinant immunocytokine, IL12-SS1 (Fv), was produced in insect cells using a baculovirus-insect cell expression system. The SS1 single-chain Fv was fused to the C terminus of the p35 subunit of IL12 through a short linker (GSADGG). The single-chain IL12-SS1 (Fv) immunocytokine bound native mesothelin proteins on malignant mesothelioma (NCI-H226) and ovarian (OVCAR-3) cells as well as recombinant mesothelin on A431/H9 cells. The immunocytokine retained sufficient bioactivity of IL12 and significantly inhibited human malignant mesothelioma (NCI-H226) grown in the peritoneal cavity of nude mice and showed comparable anti-tumor activity to that of the SS1P immunotoxin. IL12-SS1 (Fv) is the first reported immunocytokine to mesothelin-positive tumors and may be an attractive addition to mesothelin-targeted cancer therapies. PMID:24260587

  4. Novel immunocytokine IL12-SS1 (Fv inhibits mesothelioma tumor growth in nude mice.

    Directory of Open Access Journals (Sweden)

    Heungnam Kim

    Full Text Available Mesothelin is a glycosylphosphatidylinositol-anchored glycoprotein that is highly expressed on the cell surface of malignant mesothelioma. Monoclonal antibodies against mesothelin are being evaluated for the treatment of mesothelioma. Immunocytokines represent a novel class of armed antibodies. To provide an alternative approach to current mesothelin-targeted antibody therapies, we have developed a novel immunocytokine based on interleukin-12 (IL12 and the SS1 Fv specific for mesothelin. IL12 possesses potent anti-tumor activity in a wide variety of solid tumors. The newly-developed recombinant immunocytokine, IL12-SS1 (Fv, was produced in insect cells using a baculovirus-insect cell expression system. The SS1 single-chain Fv was fused to the C terminus of the p35 subunit of IL12 through a short linker (GSADGG. The single-chain IL12-SS1 (Fv immunocytokine bound native mesothelin proteins on malignant mesothelioma (NCI-H226 and ovarian (OVCAR-3 cells as well as recombinant mesothelin on A431/H9 cells. The immunocytokine retained sufficient bioactivity of IL12 and significantly inhibited human malignant mesothelioma (NCI-H226 grown in the peritoneal cavity of nude mice and showed comparable anti-tumor activity to that of the SS1P immunotoxin. IL12-SS1 (Fv is the first reported immunocytokine to mesothelin-positive tumors and may be an attractive addition to mesothelin-targeted cancer therapies.

  5. Refolded scFv Antibody Fragment against Myoglobin Shows Rapid Reaction Kinetics

    Directory of Open Access Journals (Sweden)

    Hyung-Nam Song

    2014-12-01

    Full Text Available Myoglobin is one of the early biomarkers for acute myocardial infarction. Recently, we have screened an antibody with unique rapid reaction kinetics toward human myoglobin antigen. Antibodies with rapid reaction kinetics are thought to be an early IgG form produced during early stage of in vivo immunization. We produced a recombinant scFv fragment for the premature antibody from Escherichia coli using refolding technology. The scFv gene was constructed by connection of the VH–VL sequence with a (Gly4Ser3 linker. The scFv fragment without the pelB leader sequence was expressed at a high level, but the solubility was extremely low. A high concentration of 8 M urea was used for denaturation. The dilution refolding process in the presence of arginine and the redox reagents GSH and GSSH successfully produced a soluble scFv protein. The resultant refolded scFv protein showed association and dissociation values of 9.32 × 10−4 M−1·s−1 and 6.29 × 10−3 s−1, respectively, with an affinity value exceeding 107 M−1 (kon/koff, maintaining the original rapid reaction kinetics of the premature antibody. The refolded scFv could provide a platform for protein engineering for the clinical application for diagnosis of heart disease and the development of a continuous biosensor.

  6. Refolded scFv antibody fragment against myoglobin shows rapid reaction kinetics.

    Science.gov (United States)

    Song, Hyung-Nam; Jang, Jun-Hyuck; Kim, Young-Wan; Kim, Dong-Hyung; Park, Sung-Goo; Lee, Myung Kyu; Paek, Se-Hwan; Woo, Eui-Jeon

    2014-01-01

    Myoglobin is one of the early biomarkers for acute myocardial infarction. Recently, we have screened an antibody with unique rapid reaction kinetics toward human myoglobin antigen. Antibodies with rapid reaction kinetics are thought to be an early IgG form produced during early stage of in vivo immunization. We produced a recombinant scFv fragment for the premature antibody from Escherichia coli using refolding technology. The scFv gene was constructed by connection of the V(H)-V(L) sequence with a (Gly4Ser)3 linker. The scFv fragment without the pelB leader sequence was expressed at a high level, but the solubility was extremely low. A high concentration of 8 M urea was used for denaturation. The dilution refolding process in the presence of arginine and the redox reagents GSH and GSSH successfully produced a soluble scFv protein. The resultant refolded scFv protein showed association and dissociation values of 9.32 × 10⁻⁴ M⁻¹·s⁻¹ and 6.29 × 10⁻³ s⁻¹, respectively, with an affinity value exceeding 10⁷ M⁻¹ (k(on)/k(off)), maintaining the original rapid reaction kinetics of the premature antibody. The refolded scFv could provide a platform for protein engineering for the clinical application for diagnosis of heart disease and the development of a continuous biosensor. PMID:25530617

  7. Preconceptions and Relations Used by Children in the Construction of Food Chains.

    Science.gov (United States)

    Gallegos, Leticia; And Others

    1994-01-01

    Examines the predator-prey relations and the preconceptions held by children (n=506) on the construction of food chains. Results show that the classification of herbivores and carnivores is based on children's preconceptions of size and ferocity and sheds light on the difficulties students have at higher education levels in the resolution of food…

  8. In vivo imaging of prostate cancer using an anti-PSMA scFv fragment as a probe.

    Science.gov (United States)

    Mazzocco, Claire; Fracasso, Giulio; Germain-Genevois, Coralie; Dugot-Senant, Nathalie; Figini, Mariangela; Colombatti, Marco; Grenier, Nicolas; Couillaud, Franck

    2016-01-01

    We aimed to evaluate a fluorescent-labeled single chain variable fragment (scFv) of the anti-PSMA antibody as a specific probe for the detection of prostate cancer by in vivo fluorescence imaging. An orthotopic model of prostate cancer was generated by injecting LNCaP cells into the prostate lobe. ScFvD2B, a high affinity anti-PSMA antibody fragment, was labeled using a near-infrared fluorophore to generate a specific imaging probe (X770-scFvD2B). PSMA-unrelated scFv-X770 was used as a control. Probes were injected intravenously into mice with prostate tumors and fluorescence was monitored in vivo by fluorescence molecular tomography (FMT). In vitro assays showed that X770-scFvD2B specifically bound to PSMA and was internalized in PSMA-expressing LNCaP cells. After intravenous injection, X770-scFvD2B was detected in vivo by FMT in the prostate region. On excised prostates the scFv probe co-localized with the cancer cells and was found in PSMA-expressing cells. The PSMA-unrelated scFv used as a control did not label the prostate cancer cells. Our data demonstrate that scFvD2B is a high affinity contrast agent for in vivo detection of PSMA-expressing cells in the prostate. NIR-labeled scFvD2B could thus be further developed as a clinical probe for imaging-guided targeted biopsies. PMID:26996325

  9. Not As Easy As It Seems: Automating the Construction of Lexical Chains Using Roget's Thesaurus

    CERN Document Server

    Jarmasz, Mario

    2012-01-01

    Morris and Hirst present a method of linking significant words that are about the same topic. The resulting lexical chains are a means of identifying cohesive regions in a text, with applications in many natural language processing tasks, including text summarization. The first lexical chains were constructed manually using Roget's International Thesaurus. Morris and Hirst wrote that automation would be straightforward given an electronic thesaurus. All applications so far have used WordNet to produce lexical chains, perhaps because adequate electronic versions of Roget's were not available until recently. We discuss the building of lexical chains using an electronic version of Roget's Thesaurus. We implement a variant of the original algorithm, and explain the necessary design decisions. We include a comparison with other implementations.

  10. Expression of secreted human single-chain fragment variable antibody against human amyloid beta peptide in Pichia pastoris

    Institute of Scientific and Technical Information of China (English)

    Jiong Cai; Fang Li; Shizhen Wang

    2008-01-01

    BACKGROUND: Studies have shown that monoclonal or polyclonal antibody injections ofamyloid β peptide arc effective in removing amyloid β peptide overload in the brain.OBJECTIVE: Based on successful screening of a human single-chain fragment variable antibody specific to amyloid β peptide, this paper aimed to express recombinant human single-chain variable antibody against amyloid β peptide.DESIGN, TIME AND SETTING: A single sample experiment was performed at the Department of Nuclear Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Hospital (Beijing, China) from January to July 2006.MATERIALS: Human single-chain fragment variable antibody gene against amyloid β peptide was screened from a human phage-display antibody library.METHODS: Human single-chain fragment variable antibody gene was mutated to eliminate a BamHI restriction site and cloned into a Teasy plasmid for pT-seFvconstruction, which was identified by PCR amplification and endonuclease digestion. Plasmid pT-scFvA β was cut by EcoRl and Notl endonucleases, and the antibody gene was cloned into pPIC9K plasmid to construct pPIC9K-scFvA β expression vector, which was confirmed by gene sequencing. Linearized pPICgK-scFvA β was used to transform a Pichia pastoris GS115 cell line, and the recombinant was induced by 0.5 % methanol to express human single-chain fragment variable antibody specific to amyloid β peptide.MAIN OUTCOME MEASURES: Protein electrophoresis was used to identify PCR products, gene sequencing was uscd to verify the pPIC9K-scFvA sequence, and SDS-PAGE was used to detect recombinant expression of human single-chain fragment variable antibody specific to amyloid β peptide in Pichia pastoris.RESULTS: Gene sequencing confirmed pPICgK-scFvA β orientation. Rccomhinants were obtained by lineadzed pPIC9K-scFvA β transformation. After induction with 0.5% methanol, the recombinant yeast cells secreted proteins of 33-ku size

  11. Green nanotechnology in Nordic Construction: Eco-innovation strategies and Dynamics in Nordic Window Value Chains

    DEFF Research Database (Denmark)

    Andersen, Maj Munch

    2010-01-01

    This project analyzes Nordic trends in the development and industrial uptake of green nanotechno-logy in construction. The project applies an evolutionary economic perspective in analyzing the innovation dynamics and firm strategies in the window value chains in three Nordic countries, Denmark......, Finland and Sweden. Hence the project investigates two pervasive parallel market trends: The emergence of the green market and the emergence of nanotechnology. The analysis investigates how a traditional economic sector such as the construction sector reacts to such major trends. Conclusions are multiple...... of nanotechnology in the construction sector in the Nordic countries we do find quite a high number of nanotech applications in the Nordic window chains. Eco-innovation is influencing strongly on the nanotech development. We see several examples of nano-enabled smart, multifunctional green solutions in the Nordic...

  12. Green Nanotechnology in Nordic Construction - Eco-innovation strategies and Dynamics in nordic Window Chains

    DEFF Research Database (Denmark)

    Andersen, Maj Munch; Sandén, Björn A.; Palmberg, Christopher

    This project analyzes Nordic trends in the development and industrial uptake of green nanotechno-logy in construction. The project applies an evolutionary economic perspective in analyzing the innovation dynamics and firm strategies in the window value chains in three Nordic countries, Denmark......, Finland and Sweden. Hence the project investigates two pervasive parallel market trends: The emergence of the green market and the emergence of nanotechnology. The analysis investigates how a traditional economic sector such as the construction sector reacts to such major trends. Conclusions are multiple...... of nanotechnology in the construction sector in the Nordic countries we do find quite a high number of nanotech applications in the Nordic window chains. Eco-innovation is influencing strongly on the nanotech development. We see several examples of nano-enabled smart, multifunctional green solutions in the Nordic...

  13. A humanized anti-M2 scFv shows protective in vitro activity against influenza

    Energy Technology Data Exchange (ETDEWEB)

    Bradbury, Andrew M [Los Alamos National Laboratory; Velappan, Nileena [Los Alamos National Laboratory; Schmidt, Jurgen G [Los Alamos National Laboratory

    2008-01-01

    M2 is one of the most conserved influenza proteins, and has been widely prospected as a potential universal vaccine target, with protection predominantly mediated by antibodies. In this paper we describe the creation of a humanized single chain Fv from 14C2, a potent monoclonal antibody against M2. We show that the humanized scFv demonstrates similar activity to the parental mAb: it is able to recognize M2 in its native context on cell surfaces and is able to show protective in vitro activity against influenza, and so represents a potential lead antibody candidate for universal prophylactic or therapeutic intervention in influenza.

  14. EXPERIMENTAL CHALLENGE STUDY OF FV3-LIKE RANAVIRUS INFECTION IN PREVIOUSLY FV3-LIKE RANAVIRUS INFECTED EASTERN BOX TURTLES (TERRAPENE CAROLINA CAROLINA) TO ASSESS INFECTION AND SURVIVAL.

    Science.gov (United States)

    Hausmann, Jennifer C; Wack, Allison N; Allender, Matthew C; Cranfield, Mike R; Murphy, Kevin J; Barrett, Kevin; Romero, Jennell L; Wellehan, James F X; Blum, Stella A; Zink, M Christine; Bronson, Ellen

    2015-12-01

    The Maryland Zoo in Baltimore experienced an outbreak of Frog virus-3 (FV3)-like ranavirus during the summer of 2011, during which 14 of 27 (52%) of its captive eastern box turtles (Terrapene carolina carolina) survived. To assess survival, immunity, and viral shedding, an experimental challenge study was performed in which the surviving, previously infected turtles were reinfected with the outbreak strain of FV3-like ranavirus. Seven turtles were inoculated with virus intramuscularly and four control turtles received saline intramuscularly. The turtles were monitored for 8 wk with blood and oral swabs collected for quantitative polymerase chain reaction (qPCR). During that time, one of seven (14%) inoculated turtles and none of the controls (0%) died; there was no significant difference in survival. Clinical signs of the inoculated turtles, except for the turtle that died, were mild compared to the original outbreak. Quantitative PCR for FV3-like ranavirus on blood and oral swabs was positive for all inoculated turtles and negative for all controls. The turtle that died had intracytoplasmic inclusion bodies in multiple organs. Three inoculated and two control turtles were euthanized at the end of the study. No inclusion bodies were present in any of the organs. Quantitative PCR detected FV3-like ranavirus in the spleen of a control turtle, which suggested persistence of the virus. The surviving five turtles were qPCR-negative for FV3-like ranavirus from blood and oral swabs after brumation. Quantitative PCR for Terrapene herpesvirus 1 found no association between ranavirus infection and herpesvirus loads. In conclusion, previously infected eastern box turtles can be reinfected with the same strain of FV3-like ranavirus and show mild to no clinical signs but can shed the virus from the oral cavity. PMID:26667529

  15. Production of biologically active scFv and VHH antibody fragments in Bifidobacterium longum.

    Science.gov (United States)

    Shkoporov, A N; Khokhlova, E V; Savochkin, K A; Kafarskaia, L I; Efimov, B A

    2015-06-01

    Bifidobacteria constitute a significant part of healthy intestinal microbiota in adults and infants and present a promising platform for construction of genetically modified probiotic agents for treatment of gastrointestinal disorders. In this study, three strains of Bifidobacterium longum were constructed that express and secrete biologically active single-chain antibodies against human TNF-α and Clostridium difficile exotoxin A. Anti-TNF-α scFv antibody D2E7 was produced at the level of 25 μg L(-1) in broth culture and was mostly retained in the cytoplasm, while VHH-type antibodies A20.1 and A26.8 against C. difficile exotoxin A were produced at the levels of 0.3-1 mg L(-1) and secreted very efficiently. The biological activity of both antibody types was demonstrated in the mammalian cell-based assays. Expression of A20.1 and A26.8 was also observed in vivo after intragastric administration of transformed B. longum strains to (C57/BL6 × DBA/2)F1 mice. The obtained B. longum strains may serve as prototypes for construction of novel probiotic medications against inflammatory bowel disease and C. difficile-associated disease. PMID:25994292

  16. A framework for measuring the supply chain's agility of mass construction industry in Iran

    Directory of Open Access Journals (Sweden)

    Keyvan Poloie

    2012-10-01

    Full Text Available Impotent planning system in house providing process is the result of inadequate housing management system in theoretical, empirical, and operational fields. In addition, Mass Construction industry in Iran confronts with other problems such as instability in raw material prices, unsteadiness in production and investment laws and regulations, frailty of transportation infrastructure, international sanctions and etc. Furthermore, customers’ needs, lower costs, and greater customizations lead mass producing to search for new solutions and novel producing system. Agility is offered as a strategy to enable Mass Construction associations to be maintained in the competition of constantly changing market in Iran. In such a market, previous approaches lose their capabilities in supply chain. Thus to achieve agility by Mass Construction association is the chief aim of this study. This study is descriptive-analytic and can be identified as developmental –functional considering its target. After surveying previous research literature and using experts’ opinions, we investigated final agile sub criteria of supply chain and then we used interpretive- structural modeling approach to determine the relation among sub criteria and to offer an agile supply chain model. Surveying research literature and experts’ opinions lead us to identify 8 criteria (society, government, financial, information technology, market, partnership, quality and technology and also 22 sub criteria for supply chain’s agility. Then the results were analyzed through interpretive-structural approach and relation of criteria and sub criteria and their consequence were achieved. These relations showed that government and infrastructure investment, culture, regulations and responses to social and environmental issues are the basis of agility in mass housing productions’ supply chain. This model helps supply chain managers to have strategic planning to enhance agility in supply chain

  17. THE CONSTRUCTION OF MULTITYPE CANONICAL MARKOV BRANCHING CHAINS IN RANDOM ENVIRONMENTS

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The investigation for branching processes has a long history by their strong physics background, but only a few authors have investigated the branching processes in random environments. First of all, the author introduces the concepts of the multitype canonical Markov branching chain in random environment (CMBCRE) and multitype Markov branching chain in random environment (MBCRE) and proved that CMBCRE must be MBCRE, and any MBCRE must be equivalent to another CMBCRE in distribution. The main results of this article are the construction of CMBCRE and some of its probability properties.

  18. The effects of the global financial crisis on the Australian building construction supply chain

    Directory of Open Access Journals (Sweden)

    Ram Karthikeyan Thangaraj

    2012-09-01

    Full Text Available This study involves a financial analysis of 43 publicly listed and large private companies in the building and construction supply chain from 2005 to 2010; straddling the period of the global financial crisis (GFC; and examines the impact of the GFC on the performance of these companies. The construction supply chain was divided into four sectors – material suppliers, construction companies, property developers and real estate investment trusts (REITs. The findings indicate that the impact was minimal for both material suppliers and construction companies, but especially severe for the more leveraged property developers and REITs. Building material suppliers and construction companies have benefitted substantially from the building economic stimulus package provided by the Australian government to mitigate the effects of the GFC. Decreases in the valuation of assets have, to a large extent, reduced the profitability of property developers and REITs during the GFC but these companies have recovered quickly from these adverse conditions to return to a sound financial position by the end of the 2010 financial year. The results will inform investors, managers and construction professionals in devising strategies for prudent financial management and for weathering future financial crises.

  19. The effects of N-terminal insertion into VSV-G of an scFv peptide

    Directory of Open Access Journals (Sweden)

    Piechaczyk Marc

    2006-09-01

    Full Text Available Abstract Recombinant retroviruses, including lentiviruses, are the most widely used vectors for both in vitro and in vivo stable gene transfer. However, the inability to selectively deliver transgenes into cells of interest limits the use of this technology. Due to its wide tropism, stability and ability to pseudotype a range of viral vectors, vesicular stomatitis virus G protein (VSV-G is the most commonly used pseudotyping protein. Here, we attempted to engineer this protein for targeting purposes. Chimaeric VSV-G proteins were constructed by linking a cell-directing single-chain antibody (scFv to its N-terminal. We show that the chimaeric VSV-G molecules can integrate into retroviral and lentiviral particles. HIV-1 particles pseudotyped with VSV-G linked to an scFv against human Major Histocompatibility Complex class I (MHC-I bind strongly and specifically to human cells. Also, this novel molecule preferentially drives lentiviral transduction of human cells, although the titre is considerably lower that viruses pseudotyped with VSV-G. This is likely due to the inefficient fusion activity of the modified protein. To our knowledge, this is the first report where VSV-G was successfully engineered to include a large (253 amino acids exogenous peptide and where attempts were made to change the infection profile of VSV-G pseudotyped vectors.

  20. Selection of affinity-improved neutralizing human scFv against HBV PreS1 from CDR3 VH/VL mutant library.

    Science.gov (United States)

    Chen, YanMin; Bai, Yin; Guo, XiaoChen; Wang, WenFei; Zheng, Qi; Wang, FuXiang; Sun, Dejun; Li, DeShan; Ren, GuiPing; Yin, JieChao

    2016-07-01

    A CDR3 mutant library was constructed from a previously isolated anti-HBV neutralizing Homo sapiens scFv-31 template by random mutant primers PCR. Then the library was displayed on the inner membrane surface in Escherichia coli periplasmic space. Seven scFv clones were isolated from the mutant library through three rounds of screening by flow cytometry. Competition ELISA assay indicates that isolated scFv fragments show more efficient binding ability to HBV PreS1 compared with parental scFv-31. HBV neutralization assay indicated that two clones (scFv-3 and 59) show higher neutralizing activity by blocking the HBV infection to Chang liver cells. Our method provides a new strategy for rapid screening of mutant antibody library for affinity-enhanced scFv clones and the neutralizing scFvs obtained from this study provide a potential alternative of Hepatitis B immune globulin. PMID:27255707

  1. Cloning, expression, purification, and characterization of LC-1 ScFv with GFP tag

    Institute of Scientific and Technical Information of China (English)

    LU Min; GONG Xing-guo; YU Hong; LI Jian-yong

    2005-01-01

    Total RNA was isolated from the hybridoma cell line (LC-1), which secretes anti-lung adenocarcinoma monoclonal antibody, and was transferred into cDNA. Based on the FR1 (framework region 1) and FR4 conserved regions of LC-I gene, the variable regions of heavy chain (Vh) and light chain (Vl) were amplified, and the Vh and modified Vl were connected to single chain Fv (ScFv) by SOE-PCR (splice overlap extension PCR). The modified ScFv was fused with green fluorescent protein (GFP)and introduced into E. coli JMi09. The fusion protein induced by IPTG (Isopropylthiogalactoside) was about 57000 on a 10%SDS-PAGE gel (10% Sds Polyacrylamide Gel Electrophoresis), and primarily manifested as inclusion bodies. The renatured protein purified by Ni-NTA Superflow resins showed ability to bind to antigen on SPC-A-1 lung adenocarcinoma. In addition, the induced host cells fluoresced bright green under 395 nm wavelength, which indicated that the expected protein with dual activity was expressed in the prokaryotic system. The ScFv with GFP tag used in this research can be applied as a new reagent to detect immunological dye, and provide a feasible way to detect adenocarcinoma in a clinical setting.

  2. Production of a soluble single-chain variable fragment antibody against okadaic acid and exploration of its specific binding.

    Science.gov (United States)

    He, Kuo; Zhang, Xiuyuan; Wang, Lixia; Du, Xinjun; Wei, Dong

    2016-06-15

    Okadaic acid is a lipophilic marine algal toxin commonly responsible for diarrhetic shellfish poisoning (DSP). Outbreaks of DSP have been increasing and are of worldwide public health concern; therefore, there is a growing demand for more rapid, reliable, and economical analytical methods for the detection of this toxin. In this study, anti-okadaic acid single-chain variable fragment (scFv) genes were prepared by cloning heavy and light chain genes from hybridoma cells, followed by fusion of the chains via a linker peptide. An scFv-pLIP6/GN recombinant plasmid was constructed and transformed into Escherichia coli for expression, and the target scFv was identified with IC-CLEIA (chemiluminescent enzyme immunoassay). The IC15 was 0.012 ± 0.02 μg/L, and the IC50 was 0.25 ± 0.03 μg/L. The three-dimensional structure of the scFv was simulated with computer modeling, and okadaic acid was docked to the scFv model to obtain a putative structure of the binding complex. Two predicted critical amino acids, Ser32 and Thr187, were then mutated to verify this theoretical model. Both mutants exhibited significant loss of binding activity. These results help us to understand this specific scFv-antigen binding mechanism and provide guidance for affinity maturation of the antibody in vitro. The high-affinity scFv developed here also has potential for okadaic acid toxin detection. PMID:26772159

  3. Fast conversion of scFv to Fab antibodies using type IIs restriction enzymes.

    Science.gov (United States)

    Sanmark, Hanna; Huovinen, Tuomas; Matikka, Tero; Pettersson, Tiina; Lahti, Maria; Lamminmäki, Urpo

    2015-11-01

    Single chain variable fragment (scFv) antibody libraries are widely used for developing novel bioaffinity reagents, although Fab or IgG molecules are the preferred antibody formats in many final applications. Therefore, rapid conversion methods for combining multiple DNA fragments are needed to attach constant domains to the scFv derived variable domains. In this study we describe a fast and easy cloning method for the conversion of single framework scFv fragments to Fab fragments using type IIS restriction enzymes. All cloning steps excluding plating of the Fab transformants can be done in 96 well plates and the procedure can be completed in one working day. The concept was tested by converting 69 scFv clones into Fab format on 96 well plates, which resulted in 93% success rate. The method is particularly useful as a high-throughput tool for the conversion of the chosen scFv clones into Fab molecules in order to analyze them as early as possible, as the conversion can significantly affect the binding properties of the chosen clones. PMID:26271437

  4. Important fission product nuclides identification method for simplified burnup chain construction

    International Nuclear Information System (INIS)

    A method of identifying important fission product (FP) nuclides which are included in a simplified burnup chain is proposed. This method utilizes adjoint nuclide number densities and contribution functions which quantify the importance of nuclide number densities to the target nuclear characteristics: number densities of specific nuclides after burnup. Numerical tests with light water reactor (LWR) fuel pin-cell problems reveal that this method successfully identifies important FP nuclides included in a simplified burnup chain, with which number densities of target nuclides after burnup are well reproduced. A simplified burnup chain consisting of 138 FP nuclides is constructed using this method, and its good performance for predictions of number densities of target nuclides and reactivity is demonstrated against LWR pin-cell problems and multi-cell problem including gadolinium-bearing fuel rod. (author)

  5. Constructing Agri-food Industry Input-Output Data: A Value Chain Modelling Approach

    OpenAIRE

    Xayavong, Xila; Islam, Nazrul

    2009-01-01

    Various methods can be used to construct input-output data for sectoral modelling. These methods are broadly classified in the literature as commodity based survey, non-survey and hybrid approaches. Each approach has its own strengths and weaknesses. This paper presents an alternative approach to map the flows of goods and services. The case study of generating inputoutput data for agricultural industries in Western Australia is used to show how value chain modelling can accurately and reliab...

  6. The design and construction of an electronic model of the chain reaction process

    International Nuclear Information System (INIS)

    The design and construction of an electronic model simulating the nuclear chain reaction process up to three successive neutron generations is briefly described. This model is equipped with a special sound effect. However, a tape recorder can be attached and replayed in synchronism with the display with minimum hardware modifications. In this manner, it can function as a useful educational aid. The circuit is built around easily available ttl integrated circuits. (author)

  7. Constructions of the Average Rate of Return of Pension or Investment Funds Based on Chain Indices

    Directory of Open Access Journals (Sweden)

    Jacek Białek

    2014-12-01

    Full Text Available In this paper we consider the problem of the proper construction of the average rate of return of pension (or investment funds. We refer to some economical postulates given by Gajek and Kaluszka (2000. We present, discuss and compare several measures of the average rate of return of funds. We also present alternative measures based on original chain indices. We take into consideration discrete and continuous time stochastic models.

  8. Constructions of the Average Rate of Return of Pension or Investment Funds Based on Chain Indices

    OpenAIRE

    Jacek Białek

    2014-01-01

    In this paper we consider the problem of the proper construction of the average rate of return of pension (or investment) funds. We refer to some economical postulates given by Gajek and Kaluszka (2000). We present, discuss and compare several measures of the average rate of return of funds. We also present alternative measures based on original chain indices. We take into consideration discrete and continuous time stochastic models.

  9. Erratum to "Surface display of an anti-DEC-205 single chain Fv fragment in Lactobacillus plantarum increases internalization and plasmid transfer to dendritic cells in vitro and in vivo"

    OpenAIRE

    Michon, Christophe; Kuczkowska, Katarzyna; Langella, Philippe; Eijsink, Vincent G. H.; Mathiesen, Geir

    2015-01-01

    Background: Lactic acid bacteria (LAB) are promising vehicles for delivery of a variety of medicinal compounds, including antigens and cytokines. It has also been established that Lare able to deliver cDNA to host cells. To increase the efficiency of LAB-driven DNA delivery we have constructed Lactobacillus plantarum strains targeting DEC-205, which is a receptor located at the surface of dendritic cells (DCs). The purpose was to increase uptake of bacterial cells, which could lead to improve...

  10. Production, purification, and characterization of human scFv antibodies expressed in Saccharomyces cerevisiae, Pichia pastoris, and Escherichia coli.

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Feldhaus, Jane M.; Gray, Sean A.; Siegel, Robert W.; Feldhaus, Michael J.

    2005-08-01

    Single chain (scFv) antibodies are used as affinity reagents for diagnostics, therapeutics, and proteomic analyses. The antibody discovery platform we use to identify novel antigen binders involves discovery, characterization, and production. The discovery and characterization components have previously been characterized but in order to fully utilize the capabilities of affinity reagents from our yeast surface display library, efforts were focused on developing a production component to obtain purified, soluble, and active scFvs. Instead of optimizing conditions to achieve maximum yield, efforts were focused on using a system that could quickly and easily produce and process hundreds of scFv antibodies. Heterologous protein expression in Saccharomyces cerevisiae, Pichia pastoris, and Escherichia coli were evaluated for their ability to rapidly, efficaciously, and consistently produce scFv antibodies for use in downstream proteomic applications. Following purification, the binding activity of several scFv antibodies were quantified using a novel Biacore assay. All three systems produced soluble scFv antibodies which ranged in activity from 0-99%. scFv antibody yields from Saccharomyces, Pichia, and E. coli were 1.5-4.2, 0.4-7.3, and 0.63-16.4 mg L-1 culture, respectively. For our purposes, expression in E. coli proved to be the quickest and most consistent way to obtain and characterize purified scFv for downstream applications. The E. coli expression system was also used to compare scFv production levels from the periplasm, inclusion bodies, and culture media. The E. coli production system was then used to produce variants of several scFv to determine structure function relationships.

  11. Silicio: Presente e Futuro del FV

    OpenAIRE

    VIRTUANI ALESSANDRO

    2006-01-01

    Oltre il 90% dei moduli oggi in commercio sono in Silicio, elemento che rappresenterà ancora a lungo la principale materia prima del fotovoltaico. Sono ora in fase di studio nuovi processi di produzione dei wafer di Silicio, alcuni quasi maturi per venire applicati nella produzione industriale e che potrebbero rendere il fv assai competitivo con le fonti di energia tradizionali.

  12. Information Sharing and Channel Construction of Supply Chain under Asymmetric Demand Information

    Directory of Open Access Journals (Sweden)

    Guangdong Wu

    2014-01-01

    Full Text Available Information sharing and marketing channel building have become an important problem of supply chain management theory and practice. The research of information sharing focused on traditional channel of supply chain between upstream and downstream enterprises; however, the research ignores the behavior of information sharing with potential entrants and composite structure characteristics about traditional marketing channel with the direct channel. This paper uses the model to research the effects brought about sharing demand information with potential entrants and building marketing channel, which reveals information sharing and channel building mechanism in the supply chain. The study found that the five-force model of Porter regards potential entrants only as a threat that is one-sided. When the channel competitiveness meets certain conditions, manufacturer and retailer will share demand information with potential entrants. Building composite marketing channel is the manufacturer's absolute dominant strategy. Channel construction will increase the entry barriers for potential entrants and weaken the effect of double marginalization; meanwhile, the performance of supply chain will be augmented.

  13. The Development of a Recombinant scFv Monoclonal Antibody Targeting Canine CD20 for Use in Comparative Medicine.

    Science.gov (United States)

    Jain, Saurabh; Aresu, Luca; Comazzi, Stefano; Shi, Jianguo; Worrall, Erin; Clayton, John; Humphries, William; Hemmington, Sandra; Davis, Paul; Murray, Euan; Limeneh, Asmare A; Ball, Kathryn; Ruckova, Eva; Muller, Petr; Vojtesek, Borek; Fahraeus, Robin; Argyle, David; Hupp, Ted R

    2016-01-01

    Monoclonal antibodies are leading agents for therapeutic treatment of human diseases, but are limited in use by the paucity of clinically relevant models for validation. Sporadic canine tumours mimic the features of some human equivalents. Developing canine immunotherapeutics can be an approach for modeling human disease responses. Rituximab is a pioneering agent used to treat human hematological malignancies. Biologic mimics that target canine CD20 are just being developed by the biotechnology industry. Towards a comparative canine-human model system, we have developed a novel anti-CD20 monoclonal antibody (NCD1.2) that binds both human and canine CD20. NCD1.2 has a sub-nanomolar Kd as defined by an octet red binding assay. Using FACS, NCD1.2 binds to clinically derived canine cells including B-cells in peripheral blood and in different histotypes of B-cell lymphoma. Immunohistochemical staining of canine tissues indicates that the NCD1.2 binds to membrane localized cells in Diffuse Large B-cell lymphoma, Marginal Zone Lymphoma, and other canine B-cell lymphomas. We cloned the heavy and light chains of NCD1.2 from hybridomas to determine whether active scaffolds can be acquired as future biologics tools. The VH and VL genes from the hybridomas were cloned using degenerate primers and packaged as single chains (scFv) into a phage-display library. Surprisingly, we identified two scFv (scFv-3 and scFv-7) isolated from the hybridoma with bioactivity towards CD20. The two scFv had identical VH genes but different VL genes and identical CDR3s, indicating that at least two light chain mRNAs are encoded by NCD1.2 hybridoma cells. Both scFv-3 and scFv-7 were cloned into mammalian vectors for secretion in CHO cells and the antibodies were bioactive towards recombinant CD20 protein or peptide. The scFv-3 and scFv-7 were cloned into an ADEPT-CPG2 bioconjugate vector where bioactivity was retained when expressed in bacterial systems. These data identify a recombinant anti-CD20

  14. The Development of a Recombinant scFv Monoclonal Antibody Targeting Canine CD20 for Use in Comparative Medicine

    Science.gov (United States)

    Jain, Saurabh; Aresu, Luca; Comazzi, Stefano; Shi, Jianguo; Worrall, Erin; Clayton, John; Humphries, William; Hemmington, Sandra; Davis, Paul; Murray, Euan; Limeneh, Asmare A.; Ball, Kathryn; Ruckova, Eva; Muller, Petr; Vojtesek, Borek; Fahraeus, Robin; Argyle, David; Hupp, Ted R.

    2016-01-01

    Monoclonal antibodies are leading agents for therapeutic treatment of human diseases, but are limited in use by the paucity of clinically relevant models for validation. Sporadic canine tumours mimic the features of some human equivalents. Developing canine immunotherapeutics can be an approach for modeling human disease responses. Rituximab is a pioneering agent used to treat human hematological malignancies. Biologic mimics that target canine CD20 are just being developed by the biotechnology industry. Towards a comparative canine-human model system, we have developed a novel anti-CD20 monoclonal antibody (NCD1.2) that binds both human and canine CD20. NCD1.2 has a sub-nanomolar Kd as defined by an octet red binding assay. Using FACS, NCD1.2 binds to clinically derived canine cells including B-cells in peripheral blood and in different histotypes of B-cell lymphoma. Immunohistochemical staining of canine tissues indicates that the NCD1.2 binds to membrane localized cells in Diffuse Large B-cell lymphoma, Marginal Zone Lymphoma, and other canine B-cell lymphomas. We cloned the heavy and light chains of NCD1.2 from hybridomas to determine whether active scaffolds can be acquired as future biologics tools. The VH and VL genes from the hybridomas were cloned using degenerate primers and packaged as single chains (scFv) into a phage-display library. Surprisingly, we identified two scFv (scFv-3 and scFv-7) isolated from the hybridoma with bioactivity towards CD20. The two scFv had identical VH genes but different VL genes and identical CDR3s, indicating that at least two light chain mRNAs are encoded by NCD1.2 hybridoma cells. Both scFv-3 and scFv-7 were cloned into mammalian vectors for secretion in CHO cells and the antibodies were bioactive towards recombinant CD20 protein or peptide. The scFv-3 and scFv-7 were cloned into an ADEPT-CPG2 bioconjugate vector where bioactivity was retained when expressed in bacterial systems. These data identify a recombinant anti-CD20

  15. Delineation of BmSXP antibody V-gene usage from a lymphatic filariasis based immune scFv antibody library.

    Science.gov (United States)

    Rahumatullah, Anizah; Ahmad, Azimah; Noordin, Rahmah; Lim, Theam Soon

    2015-10-01

    Phage display technology is an important tool for antibody generation or selection. This study describes the development of a scFv library and the subsequent analysis of identified monoclonal antibodies against BmSXP, a recombinant antigen for lymphatic filariasis. The immune library was generated from blood of lymphatic filariasis infected individuals. A TA based intermediary cloning approach was used to increase cloning efficiency for the library construction process. A diverse immune scFv library of 10(8) was generated. Six unique monoclonal antibodies were identified from the 50 isolated clones against BmSXP. Analysis of the clones showed a bias for the IgHV3 and Vκ1 (45.5%) and IgHV2 and Vκ3 (27.3%) gene family. The most favored J segment for light chain is IgKJ1 (45.5%). The most favored D and J segment for heavy chain are IgHD6-13 (75%) and IgHJ3 (47.7%). The information may suggest a predisposition of certain V genes in antibody responses against lymphatic filariasis. PMID:26277276

  16. HBscFv-IFNγ在毕赤酵母X33中的表达、纯化及鉴定%Expression Purification and Verification of HBscFv-IFNγ in Pichia pastoris x33

    Institute of Scientific and Technical Information of China (English)

    周世水; 王小宁

    2008-01-01

    为了有效治疗乙肝病而研究了将抗乙肝病毒表面抗原单链抗体(single-chain Fv against HBV surface antigen,HBscFv)与临床治疗乙肝常用的γ-干扰素(γ-interferon,IFNγ)连接的融合蛋白(HBscFv-IFNγ).采用重叠PCR法将基因hbscfv与ifnγ连接成hbscfv-ifnγ,再构建成多拷贝重组质粒pPICZaA/(hbscfv-ifnγ)1.2.4,然后转入巴斯德毕赤酵母X33.从中筛选出的工程菌株X4能够分泌表达目的蛋白HBscFv-IFNγ,并用SDS-PAGE.Western blotting和ELISA方法进行了初步鉴定.结果表明组成HBscFv-IFNγ的HBscFv和IFNγ仍具有生物学活性.用14F7亲合层析纯化X4的发酵液可获得纯度迭95%~98%的HBscFv-IFNγ.它可中和HBV转基因小鼠血清中27.9%的乙肝病毒表面抗原(HBV surface antigen,HbsAg),这表明HBscFv-IFNγ上的抗体能够与生物体内的HBV有效结合.可见,HBscFv-IFNγ将是一种防治乙肝病而有开发前景的靶向新药.

  17. Identification of Fusarium virguliforme FvTox1-Interacting Synthetic Peptides for Enhancing Foliar Sudden Death Syndrome Resistance in Soybean.

    Directory of Open Access Journals (Sweden)

    Bing Wang

    Full Text Available Soybean is one of the most important crops grown across the globe. In the United States, approximately 15% of the soybean yield is suppressed due to various pathogen and pests attack. Sudden death syndrome (SDS is an emerging fungal disease caused by Fusarium virguliforme. Although growing SDS resistant soybean cultivars has been the main method of controlling this disease, SDS resistance is partial and controlled by a large number of quantitative trait loci (QTL. A proteinacious toxin, FvTox1, produced by the pathogen, causes foliar SDS. Earlier, we demonstrated that expression of an anti-FvTox1 single chain variable fragment antibody resulted in reduced foliar SDS development in transgenic soybean plants. Here, we investigated if synthetic FvTox1-interacting peptides, displayed on M13 phage particles, can be identified for enhancing foliar SDS resistance in soybean. We screened three phage-display peptide libraries and discovered four classes of M13 phage clones displaying FvTox1-interacting peptides. In vitro pull-down assays and in vivo interaction assays in yeast were conducted to confirm the interaction of FvTox1 with these four synthetic peptides and their fusion-combinations. One of these peptides was able to partially neutralize the toxic effect of FvTox1 in vitro. Possible application of the synthetic peptides in engineering SDS resistance soybean cultivars is discussed.

  18. Preparation of single chain variable fragment of MG7 mAb by phage display technology

    Institute of Scientific and Technical Information of China (English)

    Zhao-Cai Yu; Jie Ding; Yong-Zhan Nie; Dai-Ming Fan; Xue-Yong Zhang

    2001-01-01

    AIM To develop the single chain variable fragment of MG7 murine anti-human gastric cancer monoclonal antibody using the phage display technology for obtaining a tumor-targeting mediator. METHODS mRNA was isolated from MG7-producing murine hybridoma cell line and converted into cDNA. The variable fragments of heavy and light chain were amplified separately and assembled into ScFv with a specially constructed DNA linker by PCR. The ScFvs DNA was ligated into the phagmid vector pCANTAB5E and the ligated sample was transformed into competent E. Coli TG1. The transformed cells were infected with M13K07 helper phage to form MG7 recombinant phage antibody library. The volume and recombinant rate of the library were evaluated by means of bacterial colony count and restriction analysis. After two rounds of panning with gastric cancer cell line KATOⅢ of highly expressing MG7binding antigen, the phage clones displaying ScFv of the antibody were selected by ELISA from the enriched phage clones. The antigen-binding affinity of the positive clone was detected by competition ELISA. HB2151 E. Coli was transfected with the positive phage clone demonstrated by competition ELISA for production of a soluble form of the MG7 ScFv. ELISA assay was used to detect the antigenbinding affinity of the soluble MG7 ScFv. Finally, the relative molecular mass of soluble MG7 ScFv was measured by SDS-PAGE. RESULTS The VH, VL and ScFv DNAs were about 340bp,320bp and 750bp, respectively. The volume of the library was up to 2 × 106 and 8 of 11 random clones were recombinants. Two phage clones could strongly compete with the original MG7 antibody for binding to the antigen expressed on KATO Ⅲ cells. Within 2 strong positive phage clones, the soluble MG7 ScFv from one clone was found to have the binding activity with KATO Ⅲ cells.SDS-PAGE showed that the relative molecular weight of soluble MG7 ScFv was 32. CONCLUSION The MG7 ScFv was successfully produced by phage antibody technology, which may

  19. 从scFv噬菌体库分离特异性的人源化抗D-dimer抗体%Isolation of specific humanized anti-D-dimer scFv fragments from scFv phage libraries

    Institute of Scientific and Technical Information of China (English)

    夏红利; 谭最; 陈德杰; 乔建国; 邱仁峰

    2011-01-01

    目的 从scFv(单链Fv)噬菌体抗体库分离出对D-dimer有特异性的人源化单克隆scFv.方法 对Tomlinson scFv噬菌体文库进行3轮淘洗,富集特异性的抗D-dimer抗体并进行ELISA 验证.通过酶联免疫检测和双脱氧终止法基因测序,获取特异性的人源化单克隆抗体.结果 3轮淘洗选择出38个抗D-dimer噬菌体抗体,酶联免疫和基因测序分析后,20个不同的全长单克隆抗D-dimer scFv噬菌体抗体被筛选出来,3轮选择后阳性克隆获取率为100%;分泌性抗体ELISA结果显示单克隆anti-D-dimer噬菌体顺利表达了抗体蛋白;5个A450值较高的单克隆中,3个显示了对D-dimer的高特异性和亲和力.结论 抗体噬菌体展示技术是分离获取人源化特异性anti-D-dimer抗体的高效快速方法.%Objective To isolate specific humanized anti-D-dimer scFv(single chain Fv) antibody from scFv phage libraries. Methods Isolate anti-D-dimer positive clones from Tomlinson I + J phage libraries by three rounds of panuing, then sequence monoclonal genes by bideoxy-mediated chain termination and express soluble scFv antibody; Pick out anti-D-dimer antibodies with high specificity and affinity by ELISA.Results After three rounds of selection from human scFv phage libraries Tomlinson I and J, 38 monclonal specific anti-D-dimer scFv fragments were selected. By polyclonal and monoclonal phage ELISA and gene sequencing, 20 different full-length monoclonal scFv phages were identified, the result of soluble scFv ELISA showed that 20 full-length monoclonal scFv were expressed smoothly. According to the result of soluble scFv ELISA, in 5 scFv antibodies with high value of A450 selected, 3 scFv antibody fragments showed high specific and affinity. Conclusion Antibody phage display was an effective, rapid method to isolate anti-D-dimer antibodies with high specificity and affinity.

  20. Functional Characteristics and Molecular Mechanism of a New scFv Antibody Against Aβ42 Oligomers and Immature Protofibrils.

    Science.gov (United States)

    Zhang, Yuan; Sun, Yuanhong; Huai, Yangyang; Zhang, Ying-Jiu

    2015-12-01

    Amyloid β peptide (Aβ42) is a major determinant of Alzheimer's disease (AD). In this study, we studied a novel single-chain variable fragment (scFv), AS, generated from an antibody library of AD patients, which recognized and bound specifically to medium-size amyloid β peptide (Aβ42) oligomers and immature protofibrils (25-55 kDa) and, more importantly, reduced their level by blocking their formation or inducing their disassembly. Consequently, scFv AS ameliorated or prevented their cytotoxicity and protected SH-SY5Y cells and primary cultured neurons in vitro from their damage in a concentration-dependent manner. Comparison of its cytotoxicity-inhibiting and cytotoxicity-neutralizing activities indicated that scFv AS displayed its protective effect on target cells mainly due to its cytotoxicity-inhibitory activity though it could also neutralize the cytotoxicity. We also found that scFv AS could efficiently cross the in vitro BBB model with a delivery efficiency of over 70% after a 60-min post-administration. The scFv AS was a monovalent antibody with an affinity constant (KD) of 5.5 × 10(-6) M and a binding threshold of 6.25 × 10(-4) μM for Aβ42 oligomers. The molecular docking simulations of Aβ42 to scFv AS revealed that scFv AS tends to approached Aβ42 oligomers and immature protofibrils mainly by their hydrophobic interaction and then drew Aβ42 molecule into the gap between VL and VH domains of scFv AS by hydrophilic interaction between scFv AS and the N-terminal region (residues 1-15) of Aβ42 and the hydrophobic interactions between scFv AS and the middle region (residues 20-33) of Aβ42. The combination of scFv AS with Aβ42 was realized likely through an induced-fit process. PMID:25330935

  1. Cloning of scFv from hybridomas using a rational strategy: Application as a receptor to sensitive detection microcystin-LR in water.

    Science.gov (United States)

    Zhang, Xiuyuan; He, Kuo; Zhao, Ruiping; Wang, Lixia; Jin, Yandan

    2016-10-01

    Single chain variable fragment (scFv), containing of heavy and light chains (VH and VL) joined by a short peptide linker, has been used widely for immunodetection. Nevertheless, cloning functional variable genes is still a bottle neck for the scFv generation technology. Here, a rational strategy for cloning and selecting variable region genes from an anti-microcystin-LR hybridoma was devised, then the functional VH and VL genes were recloned and assembled to scFv using splicing overlap extension PCR. The resulting scFv gene was recombinantly expressed as a soluble scFv-alkaline phosphatase fusion protein (scFv-AP) by vector PLIP6/GN. Then an indirect competitive chemiluminescent enzyme immunoassay (ic-CLEIA) for detection of microcystin-LR was developed. The half-maximum inhibition concentrations (IC50) and limits of detection (LODs, IC15) were 0.81 ± 0.04 μgL(-1) and 0.13 ± 0.03 μgL(-1), respectively. With the mean coefficient of variation lowing 8%, the mean recovery in intra-assay and inter-assay were 100.06% and 96.46%, The proposed strategy should be useful for generation scFv in a rapid and simple way. PMID:27380224

  2. Clinical Applications of Phage-Derived sFvs and sFv Fusion Proteins

    OpenAIRE

    K. A. Chester; Bhatia, J.; Boxer, G.; Cooke, S. P.; Flynn, A. A.; Huhalov, A.; Mayer, A.; Pedley, R. B.; Robson, L.; S. K. Sharma; Spencer, D. I. R.; Begent, R H J

    2000-01-01

    Single chain Fv antibodies (sFvs) have been produced from filamentous bacteriophage libraries obtained from immunised mice. MFE-23, the most characterised of these sFvs, is reactive with carcinoembryonic antigen (CEA), a glycoprotein that is highly expressed in colorectal adenocarcinomas. MFE-23 has been expressed in bacteria and purified in our laboratory for two clinical trials; a gamma camera imaging trial using 123I-MFE-23 and a radioimmunoguided surgery trial using 125I-MFE-23, where tum...

  3. Insilico analysis of three different tag polypeptides with dual roles in scFv antibodies.

    Science.gov (United States)

    Mohammadi, Mozafar; Nejatollahi, Foroogh; Sakhteman, Amirhossein; Zarei, Neda

    2016-08-01

    Single chain fragment variable (scFv) antibodies are composed of variable heavy (VH) and variable light (VL) domains that are joined by a polypeptide linker. Typically, [(Gly4Ser) n] sequence is used as a linker to retain the integrity of the antigen-binding domain. Due to its low immunogenicity, this sequence cannot be used as a tag for scFv detection and purification. Several evidences have shown that the addition of an N or C-terminal tag for scFv detection and purification will result in the decreased expression and binding capacity of this antibody fragment. In this study, we substituted the traditional linker (GGGGS) with His-tag, C-myc or E-tag sequences through molecular modeling. Stability and integrity of all models were assessed by molecular dynamic (MD) simulation. Based on MD simulation analysis, the model containing E-tag sequence as a linker indicated more stability compared to other molecules. The results suggest that E-tag not only can be substituted for the traditional linker, also eliminates the necessity of using additional tag for scFv detection and purification. PMID:27113782

  4. Occupational risk induced by construction of energy-production chains. Methodology and evaluation in France

    International Nuclear Information System (INIS)

    The Centre d'etude sur l'evaluation de la protection dans le domaine nucleaire has been conducting studies for several years on the comparison of health and social effects in various electricity production chains; the global results of this research are presented in paper IAEA-SM-254/51, by Fagnani et al., at this Symposium. Attention should be paid to a particular area in risk evaluation: the calculation of effects during the construction stage. The method used, the Leontieff macro-economic analysis, should be examined in detail, for it is a powerful tool whose uses go well beyond risk calculation. Moreover, it is at this stage that the richness and ambiguities of occupational risk measurement appear most clearly. As this is a major factor in the evaluation of risks generated by techniques of electricity production, it is important to show how it is calculated and, especially, how comparative evaluations have been interpreted. Three conventional production technologies are examined (PWR, oil and coal) and two solar-powered (a thermal system extrapolated from the Themis plant built in France and a photovoltaic process). The production scenarios are adapted to the French context, with the location of the installations and the origin of the fuel clearly spelled out. The results of this evaluation are striking: with the exception of coal, which is a labour-intensive industry, a very large portion of occupational risk can be traced to the construction phase (from 67% for the PWR system to 97% for the photovoltaic process). Even in total health risk (occupational and public), construction accounts for a preponderant share. In fact, this risk should be compared to average occupational risk in the economy at large, i.e. that to which workers in electricity production chains would be exposed if they worked elsewhere. This is the meaning of the 'reference level' pointed out in the study. (author)

  5. SCM and extended integration at the lower tiers of the construction supply chain: An explorative study in the Dutch construction industry

    OpenAIRE

    Pryke, S. D.; Broft, R.; Badi, S. M.

    2014-01-01

    Several studies have underlined the potential of Supply Chain Management (SCM) in meeting the formidable challenges associated with fragmentation, adversarial relationships and insufficient customer focus in the delivery of construction projects (e.g. Dainty et al., 2001; Cox and Ireland, 2002; Gadde and Dubois, 2010). However, there remains a paucity of properly documented examples of successfully implemented SCM initiatives, particularly at the lower tiers of the supply chain. This study se...

  6. Design and analysis of an intelligent public FV lighting system; Diseno y analisis de un alumbrado publico FV inteligente

    Energy Technology Data Exchange (ETDEWEB)

    Hanessian D, Ana V.; Gordon, Manuel [Universidad Autonoma Metropolitana, Unidad Azcapotzalco, Mexico, D.F. (Mexico)

    2009-07-01

    In the Mexico's National Energy Balance of 2008, it is considered that of the total of the electrical power consumption in our country, 18% is dedicated to lighting. Conscious of the necessity of saving energy in public lighting in this article is presented the design, construction and analysis of the power consumption of a public light fed with electricity of photovoltaic cells and the control of intensity on the light in inverse way of the natural light. A lamp constructed based in light emitting diodes (LEDs) is used. This has the quality of consuming very little energy that could be provided by the sun and be stored to use it at night. With this system, proven at scale, energy savings are obtained superior to 50% of the conventional one and, in relation to the commercial photovoltaic (FV) luminaries up to 30%. [Spanish] Del Balance Nacional de Energia, de Mexico, de 2008, se considera que del total del consumo de energia electrica en nuestro pais, el 18% esta dedicado a la iluminacion. Conscientes de la necesidad de ahorrar energia en alumbrado publico, en este articulo se presenta el diseno de construccion y analisis del consumo energetico de una luminaria publica alimentada con electricidad de celdas fotovoltaicas y el control de intensidad de la luz de manera inversa a la luz natural. Se utiliza un foco construido a base a los diodos emisores de luz (LEDs, por sus siglas en ingles). Estos tienen la cualidad de consumir muy poca energia que podra ser suministrada por el sol y almacenada para utilizarla en la noche. Con este sistema, probado a escala, se logran ahorros de energia superiores al 50% del convencional y, en relacion a las luminarias fotovoltaicas (FV) comerciales, hasa el 30%.

  7. Comparison of three microbial hosts for the expression of an active catalytic scFv.

    Science.gov (United States)

    Robin, Sylvain; Petrov, Kliment; Dintinger, Thierry; Kujumdzieva, Anna; Tellier, Charles; Dion, Michel

    2003-01-01

    Antibodies represent an interesting protein framework on which catalytic functions can be grafted. In previous studies, we have reported the characterization of the catalytic antibody 4B2 obtained on the basis of the "bait and switch" strategy which catalyzes two different chemical reactions: the allylic isomerization of beta,gamma-unsaturated ketones and the Kemp elimination. We have cloned the antibody 4B2 and expressed it as a single-chain Fv (scFv) fragment in different expression systems, Escherichia coli and two yeasts species, in order to elicit the most suitable system to study its catalytic activity. The scFv4B2 was secreted as an active form in the culture medium of Pichia pastoris and Kluyveromyces lactis, which led respectively to 4 and 1.3mg/l after purification. In E. coli, different strategies were investigated to increase the cytoplasmic soluble fraction, which resulted, in all cases, in the expression of a low amount of functional antibodies. By contrast, substantial amount of scFv4B2 could be purified when it was expressed as inclusion bodies (12mg/l) and submitted to an in vitro refolding process. Its catalytic activity was measured and proved to be comparable to that of the whole IgG. However, the instability of the scFv4B2 in solution prevented from an exhaustive characterization of its activity and stabilization of this protein appears to be essential before designing strategies to improve its catalytic activity. PMID:12531284

  8. Nanoparticles Modified With Tumor-targeting scFv Deliver siRNA and miRNA for Cancer Therapy

    OpenAIRE

    Chen, Yunching; Zhu, Xiaodong; Zhang, Xiaoju; Liu, Bin; Huang, Leaf

    2010-01-01

    Targeted delivery of RNA-based therapeutics for cancer therapy remains a challenge. We have developed a LPH (liposome-polycation-hyaluronic acid) nanoparticle formulation modified with tumor-targeting single-chain antibody fragment (scFv) for systemic delivery of small interfering RNA (siRNA) and microRNA (miRNA) into experimental lung metastasis of murine B16F10 melanoma. The siRNAs delivered by the scFv targeted nanoparticles efficiently downregulated the target genes (c-Myc/MDM2/VEGF) in t...

  9. Construction, expression and characterization of the engineered antibody against tumor surface antigen, p185c-erbB-2

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The c-erbB-2 proto-oncogene encodes a 185kDa protein p185, which belongs to epidermal growth factorreceptor family. Amplification of this gene has been shown to correlate with poor clinical prognosis forcertain cancer patients. The monoclonal antibody A21 which directed against p185 specifically inhibitsproliferation of tumor cells overexpressing p185, hence allows it to be a candidate for targeted therapy. Inorder to overcome several drawbacks of murine MAb, we cloned its VH and VL genes and constructed thesingle-chain Fv (scFv) through a peptide linker. The recombinant scFvA21 was expressed in Escherichiacoli and purified by the affinity column. Subsequently it was characterized by ELISA, Western blot, cellimmunohistochemistry and FACS. All these assays showed the binding activity to extracellular domain(ECD) of p185. Based on those properties of scFvA21, we further constructed the scFv-Fc fusion moleculewith a homodimer form and the recombinant product was expressed in mammalian cells. In a series ofsubsequent analysis this fusion protein showed identical antigen binding site and activity with the parentantibody. These anti-p185 engineered antibodies have promised to be further modified as a tumor targetingdrugs, with a view of application in the diagnosis and treatment of human breast cancer.

  10. The Construction of Cold-Chain Logistics Park of Agricultural Products in Sanshui from Two-stage Perspective

    Institute of Scientific and Technical Information of China (English)

    Jianhui; HUANG; Wei; WANG; Quan; LI

    2015-01-01

    This paper firstly analyzed the operation model,market positioning,market demand forecast as well as market competition and challenges,park site selection,and transportation conditions for construction of the Cold-Chain Logistics Park of Agricultural Products in Sanshui.Then,it presented the overall planning scheme for construction of the Cold-Chain Logistics Park of Agricultural Products from a progressive two-stage perspective of overall planning and stage-by-stage implementation.The first stage mainly performs the function as a transaction platform of agricultural products and meanwhile provides customers with agricultural products storage and inspection services.The second stage adds value-added services such as distribution processing,modified atmosphere storage,freezing and refrigeration,market price information distribution,E-commerce of agricultural products and personalized services.It is expected to provide references and suggestions for the construction of the Cold-Chain Logistics Park of Agricultural Products.

  11. Method for preparation of single chain antibodies

    Science.gov (United States)

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

    2012-04-03

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

  12. Antiperiodic XXZ Chains with Arbitrary Spins: Complete Eigenstate Construction by Functional Equations in Separation of Variables

    Science.gov (United States)

    Niccoli, Giuliano; Terras, Véronique

    2015-07-01

    Generic inhomogeneous integrable XXZ chains with arbitrary spins are studied by means of the quantum separation of variables (SOV) method. Within this framework, a complete description of the spectrum (eigenvalues and eigenstates) of the antiperiodic transfer matrix is derived in terms of discrete systems of equations involving the inhomogeneity parameters of the model. We show here that one can reformulate this discrete SOV characterization of the spectrum in terms of functional T - Q equations of Baxter's type, hence proving the completeness of the solutions to the associated systems of Bethe-type equations. More precisely, we consider here two such reformulations. The first one is given in terms of Q-solutions, in the form of trigonometric polynomials of a given degree , of a one-parameter family of T - Q functional equations with an extra inhomogeneous term. The second one is given in terms of Q-solutions, again in the form of trigonometric polynomials of degree but with double period, of Baxter's usual (i.e., without extra term) T - Q functional equation. In both cases, we prove the precise equivalence of the discrete SOV characterization of the transfer matrix spectrum with the characterization following from the consideration of the particular class of Q-solutions of the functional T - Q equation: to each transfer matrix eigenvalue corresponds exactly one such Q-solution and vice versa, and this Q-solution can be used to construct the corresponding eigenstate.

  13. First steps toward the construction of a hyperphase diagram that covers different classes of short polymer chains

    Science.gov (United States)

    Sabeur, Sid

    2014-06-01

    We present the results of a multicanonical Monte Carlo study of flexible and wormlike polymer chains, where we investigate how the polymer structures observed during the simulations, mainly coil, liquid, and crystalline structures, can help to construct a hyperphase diagram that covers different polymer classes according to their thermodynamic behavior.

  14. Radiosensitization and growth inhibition of cancer cells mediated by an scFv antibody gene against DNA-PKcs in vitro and in vivo

    Directory of Open Access Journals (Sweden)

    Du Li

    2010-08-01

    Full Text Available Abstract Background Overexpression of DNA-dependent protein kinase catalytic subunit (DNA-PKcs is commonly occurred in cancers and causes radioresistance and poor prognosis. In present study, the single-chain variable antibody fragments (scFv targeting DNA-PKcs was developed for the application of radiosensitization in vitro and in vivo. A humanized semisynthetic scFv library and the phage-display antibodies technology were employed to screen DNA-PKcs scFv antibody. Methods DNA-PKcs epitopes were predicted and cloned. A humanized semisynthetic scFv library and the phage-display antibodies technology were employed to screen DNA-PKcs scFv antibody. DNA damage repair was analyzed by comet assay and immunofluorescence detection of γH2AX foci. The radiosensitization in vivo was determined on Balb/c athymic mice transplanted tumours of HeLa cells. Results Four epitopes of DNA-PKcs have been predicted and expressed as the antigens, and a specific human anti-DNA-PKcs scFv antibody gene, anti-DPK3-scFv, was obtained by screening the phage antibody library using the DNA-PKcs peptide DPK3. The specificity of anti-DPK3-scFv was verified, in vitro. Transfection of HeLa cells with the anti-DPK3-scFv gene resulted in an increased sensitivity to IR, decreased repair capability of DNA double-strand breaks (DSB detected by comet assay and immunofluorescence detection of γH2AX foci. Moreover, the kinase activity of DNA-PKcs was inhibited by anti-DPK3-scFv, which was displayed by the decreased phosphorylation levels of its target Akt/S473 and the autophosphorylation of DNA-PKcs on S2056 induced by radiation. Measurement of the growth and apoptosis rates showed that anti-DPK3-scFv enhanced the sensitivity of tumours transplanted in Balb/c athymic mice to radiation therapy. Conclusion The antiproliferation and radiosensitizing effects of anti-DPK3-scFv via targeting DNA-PKcs make it very appealing for the development as a novel biological radiosensitizer for cancer

  15. Radiosensitization and growth inhibition of cancer cells mediated by an scFv antibody gene against DNA-PKcs in vitro and in vivo

    International Nuclear Information System (INIS)

    Overexpression of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is commonly occurred in cancers and causes radioresistance and poor prognosis. In present study, the single-chain variable antibody fragments (scFv) targeting DNA-PKcs was developed for the application of radiosensitization in vitro and in vivo. A humanized semisynthetic scFv library and the phage-display antibodies technology were employed to screen DNA-PKcs scFv antibody. DNA-PKcs epitopes were predicted and cloned. A humanized semisynthetic scFv library and the phage-display antibodies technology were employed to screen DNA-PKcs scFv antibody. DNA damage repair was analyzed by comet assay and immunofluorescence detection of γH2AX foci. The radiosensitization in vivo was determined on Balb/c athymic mice transplanted tumours of HeLa cells. Four epitopes of DNA-PKcs have been predicted and expressed as the antigens, and a specific human anti-DNA-PKcs scFv antibody gene, anti-DPK3-scFv, was obtained by screening the phage antibody library using the DNA-PKcs peptide DPK3. The specificity of anti-DPK3-scFv was verified, in vitro. Transfection of HeLa cells with the anti-DPK3-scFv gene resulted in an increased sensitivity to IR, decreased repair capability of DNA double-strand breaks (DSB) detected by comet assay and immunofluorescence detection of γH2AX foci. Moreover, the kinase activity of DNA-PKcs was inhibited by anti-DPK3-scFv, which was displayed by the decreased phosphorylation levels of its target Akt/S473 and the autophosphorylation of DNA-PKcs on S2056 induced by radiation. Measurement of the growth and apoptosis rates showed that anti-DPK3-scFv enhanced the sensitivity of tumours transplanted in Balb/c athymic mice to radiation therapy. The antiproliferation and radiosensitizing effects of anti-DPK3-scFv via targeting DNA-PKcs make it very appealing for the development as a novel biological radiosensitizer for cancer therapeutic potential

  16. Value Chain Analysis of Construction Enterprise in China%国内建筑施工企业的价值链分析

    Institute of Scientific and Technical Information of China (English)

    邓正位; 叶敏

    2011-01-01

    在分析建筑行业现状的基础上,结合价值链理论和多年的建筑施工经验,构建施工企业的价值链系统,系统介绍施工企业如何对企业的行业价值链、内部价值链和相关单位价值链进行分析,最后对如何提高企业价值链管理水平提出建议.%Based on current situation of construction industry, value chain theory and building construction experience, this paper constructs value chain system of construction enterprise, presents how to analyze industry value chain. intemal value chain and related units value chain. Finally, some suggestions are put forward to enhance value chain management for construction enterprise.

  17. Screening, expression, and characterization of an anti-human oxidized low-density lipoprotein single-chain variable fragment.

    Science.gov (United States)

    Kumano-Kuramochi, Miyuki; Fujimura, Takashi; Komba, Shiro; Maeda-Yamamoto, Mari; Machida, Sachiko

    2016-09-01

    Increased levels of oxidized low-density lipoprotein (OxLDL) in the blood circulation are correlated with atherosclerosis. Monoclonal antibody-based detection systems have been reported for OxLDL. We identified novel single-chain variable fragments (scFvs) having affinity for human OxLDL and related ligands. We constructed an scFv library from nonimmunized human spleen mRNA. Two types (γ+κ and μ+λ) of scFv phage libraries were enriched by biopanning, and five scFv clones with affinity for OxLDL were identified. The γκ5 scFv, which showed the highest affinity for OxLDL, was cloned into pET-22b(+) and expressed in Escherichia coli BL21(DE3). γκ5, expressed as an inclusion body in BL21(DE3), was refolded and purified. The specificity and sensitivity of γκ5 were analyzed using enzyme-linked immunosorbent assays (ELISAs). The γκ5 scFv showed affinity for OxLDL and acetylated LDL. The sensitivity of γκ5 to low concentrations (1-2 μg/mL) of OxLDL was higher than that to AcLDL and LDL. Finally, we developed a sandwich ELISA using γκ5 and CTLD14 (a lectin-like OxLDL receptor-1 ligand recognition region), which allowed specific detection of OxLDL at a level below 0.1 μg/mL. Our results indicated that the γκ5 scFv was a promising molecule for the detection of modified LDL at very low concentrations. PMID:27038672

  18. A compact phage display human scFv library for selection of antibodies to a wide variety of antigens

    Directory of Open Access Journals (Sweden)

    Kristensen Peter

    2009-01-01

    Full Text Available Abstract Background Phage display technology is a powerful new tool for making antibodies outside the immune system, thus avoiding the use of experimental animals. In the early days, it was postulated that this technique would eventually replace hybridoma technology and animal immunisations. However, since this technology emerged more than 20 years ago, there have only been a handful reports on the construction and application of phage display antibody libraries world-wide. Results Here we report the simplest and highly efficient method for the construction of a highly useful human single chain variable fragment (scFv library. The least number of oligonucleotide primers, electroporations and ligation reactions were used to generate a library of 1.5 × 108 individual clones, without generation of sub-libraries. All possible combinations of heavy and light chains, among all immunoglobulin isotypes, were included by using a mixture of primers and overlapping extension PCR. The key difference from other similar libraries was the highest diversity of variable gene repertoires, which was derived from 140 non-immunized human donors. A wide variety of antigens were successfully used to affinity select specific binders. These included pure recombinant proteins, a hapten and complex antigens such as viral coat proteins, crude snake venom and cancer cell surface antigens. In particular, we were able to use standard bio-panning method to isolate antibody that can bind to soluble Aflatoxin B1, when using BSA-conjugated toxin as a target, as demonstrated by inhibition ELISA. Conclusion These results suggested that by using an optimized protocol and very high repertoire diversity, a compact and efficient phage antibody library can be generated. This advanced method could be adopted by any molecular biology laboratory to generate both naïve or immunized libraries for particular targets as well as for high-throughput applications.

  19. Non-immune binding of human protein Fv to immunoglobulins from various mammalian and non-mammalian species.

    Science.gov (United States)

    Bouvet, J P; Pires, R; Charlemagne, J; Pillot, J; Iscaki, S

    1991-10-01

    Reactivity of the secretory protein Fv with immunoglobulins (Ig) from various species of vertebrates was investigated. Binding was observed throughout all taxonomic classes: mammalian, avian, reptilian, amphibian and fish. Contrasting with this wide spectrum, no significant binding was found with most mammalian ungulates, such as horse (Perissodactyl), cow, sheep and goat (Artiodactyls). Nevertheless, disruption of the hydrogen bonds of Ig allowed these non-reactive molecules to bind. Such a conserved reactivity during evolution, and our previous data on the effect of the cleavage of the intra-chain disulphide bonds, suggest that protein Fv recognizes a discontinuous framework structure involving both the FR1 and FR3 regions in the variable domain of the heavy chain of Ig. PMID:1925412

  20. The role of the supply chain in the elimination and reduction of construction rework and defects: an action research approach

    OpenAIRE

    Taggart, Martin; Koskela, Lauri; Rooke, John

    2014-01-01

    Since 2007, Ireland has suffered a circa 80% reduction in construction output. This has resulted in bankruptcy, unemployment and bad debt. Contractors have attached greater emphasis to production efficiency and cost reduction as a means of survival. An action research (AR) strategy was used to improve processes adopted by a small/medium enterprise (SME) contractor for the control of defects in its supply chain. It is conservatively estimated that rework, typically, accounts for circa 5% of to...

  1. Gladiolus plants transformed with single-chain variable fragment antibodies to Cucumber mosaic virus

    Science.gov (United States)

    Transgenic plants of Gladiolus ‘Peter Pears’ or ‘Jenny Lee’ were developed that contain single-chain variable fragments (scFv) to Cucumber mosaic virus (CMV) subgroup I or II. The CMV subgroup I heavy and light chain scFv fragments were placed under control of either the duplicated CaMV 35S or suga...

  2. Information Sharing and Channel Construction of Supply Chain under Asymmetric Demand Information

    OpenAIRE

    Guangdong Wu; Qingshan Kong; Jian-gang Shi; Hamid Reza Karimi; Wei Zhang

    2014-01-01

    Information sharing and marketing channel building have become an important problem of supply chain management theory and practice. The research of information sharing focused on traditional channel of supply chain between upstream and downstream enterprises; however, the research ignores the behavior of information sharing with potential entrants and composite structure characteristics about traditional marketing channel with the direct channel. This paper uses the model to research the eff...

  3. Click ‘Like’ and Post It on Your Wall! Chain Posts on Facebook – Identity Construction and Values

    Directory of Open Access Journals (Sweden)

    Piret Voolaid

    2013-04-01

    Full Text Available The article focuses on chain posts that were collected in the years 2010–2012 and spread predominantly among girls of ten to twelve on Facebook (facebook.com – a social network that has a membership of over 450,000 in Estonia. The source material comprising approximately 220 texts is similar by form and content to chain letters known from earlier tradition; yet, the web environment with its specific technical structure allows the texts to turn into a peculiar Facebook-like phenomenon.The author takes a closer interest in the changes in form adapted to chain letters as a genre in Facebook environment as well as thematic categories of these letters. The analysis of the epistolary cultural phenomenon focuses on the socio-folkloric nature of texts with its communicative and socio-cultural aspects. The main focus is on how socio-cultural environment influences changes in the genre, what kind of global and local impacts occur in Estonian-language chain posts and how this everyday genre reflects the realities of the era and the values intrinsic to this age group. The levels of personal and collective identity construction of chain posters as a special age group have been analysed against the identity motivation theory known from social psychology.

  4. A chimera of green fluorescent protein with single chain variable fragment antibody against ginsenosides for fluorescence-linked immunosorbent assay.

    Science.gov (United States)

    Sakamoto, Seiichi; Tanizaki, Yusuke; Pongkitwitoon, Benyakan; Tanaka, Hiroyuki; Morimoto, Satoshi

    2011-05-01

    A chimera of green fluorescent protein extracted from Aequorea coerulescens (AcGFP), a mutant that has been codon optimized for mammalian expression, with single-chain variable fragment (scFv) antibody against ginsenoside Re (GRe-scFv), named fluobody, has been successfully expressed in Escherichia coli (E. coli) to develop simple, speedy, and sensitive fluorescence-linked immunosorbent assay (FLISA). Two chimera proteins were constructed to contain GRe-scFv at the C-terminus of AcGFP (C-fluobody) and at the N-terminus of AcGFP (N-fluobody). These fluobodies were then purified by ion metal affinity chromatography and refolded by stepwise dialysis. The characterization of both fluobodies revealed that C-fluobody was found to be appropriate probe for FLISA as compare with N-fluobody. Furthermore, improvement of limit of detection (LOD) was observed in FLISA using C-fluobody (10 ng/mL) due to its strong fluorescence intensity of AcGFP compared with conventional enzyme-linked immunosorbent assay (ELISA) using parental monoclonal antibody against ginsenoside Re (G-Re), MAb-4G10 (100 ng/mL). Since some steps required in ELISA can be avoided in this present FLISA, speedy and sensitive immunoassay also could be performed using fluobody instead of monoclonal antibody and scFv. PMID:21277981

  5. Design and construction of a new human naïve single-chain fragment variable antibody library, IORISS1.

    Science.gov (United States)

    Pasello, Michela; Zamboni, Silvia; Mallano, Alessandra; Flego, Michela; Picci, Piero; Cianfriglia, Maurizio; Scotlandi, Katia

    2016-04-20

    Human monoclonal antibodies are a powerful tool with increasingly successful exploitations and the single chain fragment variable format can be considered the building block for the implementation of more complex and effective antibody-based constructs. Phage display is one of the best and most efficient methods to isolate human antibodies selected from an efficient and variable phage display library. We report a method for the construction of a human naïve single-chain variable fragment library, termed IORISS1. Many different sets of oligonucleotide primers as well as optimized electroporation and ligation reactions were used to generate this library of 1.2×10(9) individual clones. The key difference is the diversity of variable gene templates, which was derived from only 15 non-immunized human donors. The method described here, was used to make a new human naïve single-chain fragment variable phage display library that represents a valuable source of diverse antibodies that can be used as research reagents or as a starting point for the development of therapeutics. Using biopanning, we determined the ability of IORISS1 to yield antibodies. The results we obtained suggest that, by using an optimized protocol, an efficient phage antibody library can be generated. PMID:26945728

  6. Supply chain partnership in construction a field study on project team level factors

    NARCIS (Netherlands)

    Koolwijk , J.S.J.; Van Oel, C.J.; Wamelink , J.W.F.

    2015-01-01

    People and their relationship are at the heart of supply chain partnerships, however there is a lack of qualitative studies focusing on how integrated relationships may be developed. Therefore, the purpose of this study was to conduct field research to deepen our understanding of team level variable

  7. Improving supply chain management in construction: what can be learned from the aerospace industry?

    NARCIS (Netherlands)

    Voordijk, H.; Vrijhoef, R.; Greenwood, D.J.

    2003-01-01

    In order to provide for controllable delivery, reliable lead times and efficient customer response, lean manufacturing and platform assembly practices play an important role in supply chains in the aerospace industry. The adoption of lean manufacturing practices ensures an efficient delivery of prod

  8. 2D warp-and-woof interwoven networks constructed by helical chains with different chirality.

    Science.gov (United States)

    Feng, Yuhua; Guo, Yang; OuYang, Yan; Liu, Zhanquan; Liao, Daizheng; Cheng, Peng; Yan, Shiping; Jiang, Zonghui

    2007-09-21

    Two unprecedented 2D entangled layers of warp-and-woof threads interwoven by left- and right-handed helical chains, {[Mn(salen)Au(CN)2]4(H2O)}n (salen = N,N'-ethylenebis(salicylideneaminato)) and {Mn(acacen)Ag(CN)2}n (acacen = N,N'-ethylenebis(acetylacetonylideneiminate)) 2, have been synthesized and characterized. PMID:17728880

  9. Modularity in supply chains: a multiple case study in the construction industry

    NARCIS (Netherlands)

    Voordijk, Hans; Meijboom, Bert; Haan, de Job

    2006-01-01

    Purpose – The objective of this study is to assess the applicability of Fine's three-dimensional modularity concept as a tool to describe and to analyze the alignment of product, process, and supply chain architectures. Fine claims that the degree of modularity in the final output product has a one-

  10. Hydrothermal synthesis, crystal structure and properties of a novel chain coordination polymer constructed by tetrafunctional phosphonate anions and cobalt ions

    International Nuclear Information System (INIS)

    A novel cobalt phosphonate, [Co(HL)(H2O)3]n (1) (L=N(CH2PO3H)33−) has been synthesized by hydrothermal reaction at 150 °C and structurally characterized by X-ray diffraction, infrared spectroscopy, elemental and thermogravimetric analysis. Complex 1 features a 1D chain structure with double-channel built from CoO6 octahedra bridged together by the phosphonate groups. Each cobalt ion is octahedrally coordinated by three phosphonate oxygen atoms and three water molecules. The coordinated water molecules can form the hydrogen bonds with the phosphonate oxygen atoms to link the 1D chains, building a 2D layered structure, further resulting in a 3D network. The luminescence spectrum indicates an emission maximum at 435 nm. The magnetic susceptibility curve exhibits a dominant antiferromagnetic behavior with a weakly ferromagnetic component at low temperatures. - Graphical abstract: The connectivity between cobalt ions and the ligands results in a chain structure with a 1D double-channel structure, which is constructed by A-type subrings and B-type subrings. - Highlights: • The tetrafunctional phosphonate ligand was used as the ligand. • A novel chain structure can be formed by A-type rings and B-type rings. • Two types of rings can form a 1D double-channel structure, along the c-axis

  11. Hydrothermal synthesis, crystal structure and properties of a novel chain coordination polymer constructed by tetrafunctional phosphonate anions and cobalt ions

    Energy Technology Data Exchange (ETDEWEB)

    Guan, Lei, E-mail: gl_coord@163.com [School of Chemistry and Materials Science, Liaoning University of Petroleum and Chemical Engineering, Fushun 113001 (China); Wang, Ying [Center of Experiment, College of Chemistry, Chemical Engineering and Environmental Engineering, Liaoning University of Petroleum and Chemical Engineering, Fushun 113001 (China)

    2015-08-15

    A novel cobalt phosphonate, [Co(HL)(H{sub 2}O){sub 3}]{sub n} (1) (L=N(CH{sub 2}PO{sub 3}H){sub 3}{sup 3−}) has been synthesized by hydrothermal reaction at 150 °C and structurally characterized by X-ray diffraction, infrared spectroscopy, elemental and thermogravimetric analysis. Complex 1 features a 1D chain structure with double-channel built from CoO{sub 6} octahedra bridged together by the phosphonate groups. Each cobalt ion is octahedrally coordinated by three phosphonate oxygen atoms and three water molecules. The coordinated water molecules can form the hydrogen bonds with the phosphonate oxygen atoms to link the 1D chains, building a 2D layered structure, further resulting in a 3D network. The luminescence spectrum indicates an emission maximum at 435 nm. The magnetic susceptibility curve exhibits a dominant antiferromagnetic behavior with a weakly ferromagnetic component at low temperatures. - Graphical abstract: The connectivity between cobalt ions and the ligands results in a chain structure with a 1D double-channel structure, which is constructed by A-type subrings and B-type subrings. - Highlights: • The tetrafunctional phosphonate ligand was used as the ligand. • A novel chain structure can be formed by A-type rings and B-type rings. • Two types of rings can form a 1D double-channel structure, along the c-axis.

  12. Research on the Construction of Project Critical Chain Model in South-to-North Water Diversion Project with Multi-Resource Constraints Based on Portfolio Technology

    Directory of Open Access Journals (Sweden)

    Jing-chun FENG

    2011-06-01

    Full Text Available Recently, the critical chain study has become the hot spot within the project management research field, so as the construction of it with multi-resource constraints become the new research subject. Referring to System Analysis Theory and Project Portfolio Theory, this paper discusses the creation of project portfolio based on project similarity, and definition of priority in multi-resource allocation based on quantitative analysis. Then according to the theory on critical chain construction, the author made relevant research and proposed the construction model with the multi-resource constraints, which to be applied to the critical chain construction of A-bid section in South-to-North Water Diversion Project. And necessary contrast analysis with the Comprehensive Treatment Construction Method and Aggressive Treatment Construction Method also has been made within this paper.

  13. Construction and Design of Post-Tensioned Pearl-Chain Bridges using SL-Technology

    DEFF Research Database (Denmark)

    Halding, Philip Skov

    2016-01-01

    a filling layer of lower stiffness above the arches to level the road surface. The present Ph.D. thesis is part of a larger development project about Pearl‐Chain Bridges funded by Innovationsfonden. The project also included another Ph.D. study about the developed materials used in Pearl‐Chain Bridges......, and numerical modelling. Despite of high levels of normal force in PC‐Bridges, the result showed that a Mesnager inspired hinge type had a response similar to what was predicted in the literature. A specially designed saddle bearing also had elastic and plastic rotational resistance, but this hinge type......) A test to fracture to observe the ductility in the system, and fracture type. The collapse occurred after two plastic hinges were formed in the 3/8, and 5/8 points of the span. Several warnings signs were observed when approaching the maximum loading....

  14. Spontaneous construction of photoactive hollow TiO2 microspheres and chains

    International Nuclear Information System (INIS)

    Discrete and chain-like aggregates of well-defined hollow TiO2 microspheres are reproducibly synthesized in high yield by a modified fluoride-mediated self-transformation strategy using urea as a base catalyst. The shell walls are composed of agglomerated polyhedral anatase nanocrystals, and exhibit hierarchical porosity. The addition of urea tunes the nucleation dynamics and surface states of the elementary TiO2 building blocks, which together promote the formation of metastable solid microparticles of uniform size that subsequently transform into morphologically invariant hollow microspheres and chain-like aggregates. The high surface area, bimodal mesoporosity of the shell walls, and increased band gap of the hollow TiO2 microspheres give rise to increases in photocatalytic activity when compared with anatase nanoparticles and aggregates prepared in the absence of urea.

  15. Construction of a recombinant bacterial plasmid containing DNA sequences for a mouse embryonic globin chain.

    Science.gov (United States)

    Fantoni, A; Bozzoni, I; Ullu, E; Farace, M G

    1979-01-01

    Messenger RNAs for mouse embryonic globins were purified from yolk sac derived eyrthroid cells in mouse fetuses. Double stranded DNAs complementary to these messengers were synthesized and blunt end ligated to a EcoRI digested and DNA polymerase I repaired pBR322 plasmid. Of the ampicillin resistant transformants, one contained a plasmid with globin-specific cDNA. The inserted sequence is about 350 base pairs long. It contains one restriction site for EcoRI and one restriction site for HinfI about 170 and 80 base pairs from one end. The insert is not cleaved by HindIII, HindII, BamHI, PstI, SalI, AvaI, TaqI, HpaII, BglI. A mixture of purified messengers coding for alpha chains and for x, y and z embryonic chains was incubated with the recombinant plasmid and the hybridized messenger was translated in a mRNA depleted reticulocyte lysate protein synthesizing system. The product of translation was identified as a z chain by carboxymethylcellulose cromatography. The recombinant plasmid is named "pBR322-egz" after embryonic globin z. Images PMID:493112

  16. Construction of a recombinant bacterial plasmid containing DNA sequences for a mouse embryonic globin chain.

    Science.gov (United States)

    Fantoni, A; Bozzoni, I; Ullu, E; Farace, M G

    1979-08-10

    Messenger RNAs for mouse embryonic globins were purified from yolk sac derived eyrthroid cells in mouse fetuses. Double stranded DNAs complementary to these messengers were synthesized and blunt end ligated to a EcoRI digested and DNA polymerase I repaired pBR322 plasmid. Of the ampicillin resistant transformants, one contained a plasmid with globin-specific cDNA. The inserted sequence is about 350 base pairs long. It contains one restriction site for EcoRI and one restriction site for HinfI about 170 and 80 base pairs from one end. The insert is not cleaved by HindIII, HindII, BamHI, PstI, SalI, AvaI, TaqI, HpaII, BglI. A mixture of purified messengers coding for alpha chains and for x, y and z embryonic chains was incubated with the recombinant plasmid and the hybridized messenger was translated in a mRNA depleted reticulocyte lysate protein synthesizing system. The product of translation was identified as a z chain by carboxymethylcellulose cromatography. The recombinant plasmid is named "pBR322-egz" after embryonic globin z. PMID:493112

  17. Engineering an Anti-Transferrin Receptor ScFv for pH-Sensitive Binding Leads to Increased Intracellular Accumulation

    OpenAIRE

    Benjamin J Tillotson; Goulatis, Loukas I.; Parenti, Isabelle; Duxbury, Elizabeth; Shusta, Eric V.

    2015-01-01

    The equilibrium binding affinity of receptor-ligand or antibody-antigen pairs may be modulated by protonation of histidine side-chains, and such pH-dependent mechanisms play important roles in biological systems, affecting molecular uptake and trafficking. Here, we aimed to manipulate cellular transport of single-chain antibodies (scFvs) against the transferrin receptor (TfR) by engineering pH-dependent antigen binding. An anti-TfR scFv was subjected to histidine saturation mutagenesis of a s...

  18. Use of 2D NMR, protein engineering, and molecular modeling to study the hapten-binding site of an antibody Fv fragment against 2-phenyloxazolone

    International Nuclear Information System (INIS)

    Two-dimensional (2D) 1H NMR spectroscopy was used to study the hapten-binding site of a recombinant antibody Fv fragment expressed in Escherichia coli. Point mutations of residues in the CDR loops of the Fv fragment were designed in order to investigate their influence on hapten binding and to make site-specific assignments of aromatic NMR proton signals. Two tyrosines giving NOEs to the ligand 2-phenyloxazolone were identified, residue 33 in CDR1 of the heavy chain and residue 32 in CDR1 of the light chain. The benzyl portion of 2-phenyloxazolone is located between these two residues. The binding site is close to the surface of the Fv fragment. Comparison with a different anti-2-phenyloxazolone antibody, the crystal structure of which has recently been solved, shows that the general location of the hapten-binding site in both antibodies is similar. However, in the crystallographically solved antibody, the hapten is bound farther from the surface in a pocket created by a short CDR3 loop of the heavy chain. In the binding site identified in the Fv fragment studied in this report, this space is probably filled by the extra seven residues of the CDR3

  19. Construction of a rationally designed antibody platform for sequencing-assisted selection.

    Science.gov (United States)

    Larman, H Benjamin; Xu, George Jing; Pavlova, Natalya N; Elledge, Stephen J

    2012-11-01

    Antibody discovery platforms have become an important source of both therapeutic biomolecules and research reagents. Massively parallel DNA sequencing can be used to assist antibody selection by comprehensively monitoring libraries during selection, thus greatly expanding the power of these systems. We have therefore constructed a rationally designed, fully defined single-chain variable fragment (scFv) library and analysis platform optimized for analysis with short-read deep sequencing. Sequence-defined oligonucleotide libraries encoding three complementarity-determining regions (L3 from the light chain, H2 and H3 from the heavy chain) were synthesized on a programmable microarray and combinatorially cloned into a single scFv framework for molecular display. Our unique complementarity-determining region sequence design optimizes for protein binding by utilizing a hidden Markov model that was trained on all antibody-antigen cocrystal structures in the Protein Data Bank. The resultant ~10(12)-member library was produced in ribosome-display format, and comprehensively analyzed over four rounds of antigen selections by multiplex paired-end Illumina sequencing. The hidden Markov model scFv library generated multiple binders against an emerging cancer antigen and is the basis for a next-generation antibody production platform. PMID:23064642

  20. Optimizing the supply chain for passive materials in network construction projects of a telecom operator

    OpenAIRE

    Bergman, Leo Bergman

    2016-01-01

    Manufacturing Planning and Control (MPC) is a hierarchical manufacturing paradigm that has not been studied much in a project-oriented production environment. The few articles that have covered the field have come to a conclusion that it is either not feasible at all or that some aspects of MPC can be applied also in project business. In this thesis, MPC is tested in a project environment by conducting a case study on the supply chain of passive materials in optical network projects of a ...

  1. Brief Communication: Earthquake sequencing: analysis of time series constructed from the Markov chain model

    Science.gov (United States)

    Cavers, M. S.; Vasudevan, K.

    2015-10-01

    Directed graph representation of a Markov chain model to study global earthquake sequencing leads to a time series of state-to-state transition probabilities that includes the spatio-temporally linked recurrent events in the record-breaking sense. A state refers to a configuration comprised of zones with either the occurrence or non-occurrence of an earthquake in each zone in a pre-determined time interval. Since the time series is derived from non-linear and non-stationary earthquake sequencing, we use known analysis methods to glean new information. We apply decomposition procedures such as ensemble empirical mode decomposition (EEMD) to study the state-to-state fluctuations in each of the intrinsic mode functions. We subject the intrinsic mode functions, derived from the time series using the EEMD, to a detailed analysis to draw information content of the time series. Also, we investigate the influence of random noise on the data-driven state-to-state transition probabilities. We consider a second aspect of earthquake sequencing that is closely tied to its time-correlative behaviour. Here, we extend the Fano factor and Allan factor analysis to the time series of state-to-state transition frequencies of a Markov chain. Our results support not only the usefulness of the intrinsic mode functions in understanding the time series but also the presence of power-law behaviour exemplified by the Fano factor and the Allan factor.

  2. Diagnostic potential of recombinant scFv antibodies generated against hemagglutinin protein of influenza A virus

    Directory of Open Access Journals (Sweden)

    Roopali eRajput

    2015-09-01

    Full Text Available Human influenza A viruses have been the cause of enormous socio-economic losses worldwide. In order to combat such a notorious pathogen, hemagglutinin protein (HA has been a preferred target for generation of neutralizing-antibodies, as potent therapeutic/ diagnostic agents. In the present study, recombinant anti-HA single chain variable fragment (scFv antibodies were constructed using the phage display technology to aid in diagnosis and treatment of human influenza A virus infections. Spleen cells of mice hyper-immunized with A/New Caledonia/20/99 (H1N1 virus were used as the source for recombinant antibody (rAb production. The antigen-binding phages were quantified after 6 rounds of bio-panning against A/New Caledonia/20/99 (H1N1, A/California/07/2009 (H1N1-like, or A/Udorn/307/72(H3N2 viruses. The phage yield was maximum for the A/New Caledonia/20/99 (H1N1, however, considerable cross-reactivity was observed for the other virus strains as well. The HA-specific polyclonal rAb preparation was subjected to selection of single clones for identification of high reactive relatively conserved epitopes. The high affinity rAbs were tested against certain known conserved HA epitopes by peptide ELISA. Three recombinant mAbs showed reactivity with both the H1N1 strains and one (C5 showed binding with all the three viral strains. The C5 antibody was thus used for development of an ELISA test for diagnosis of influenza virus infection. Based on the sample size in the current analysis, the ELISA test demonstrated 83.9% sensitivity and 100% specificity. Thus, the ELISA, developed in our study, may prove as a cheaper alternative to the presently used real time RT-PCR test for detection of human influenza A viruses in clinical specimens, which will be beneficial, especially in the developing countries. Since, the two antibodies identified in this study are reactive to conserved HA epitopes; these may prove as potential therapeutic agents as well.

  3. Need of a consistent and convenient nucleus identification in ENDF files for the automatic construction of the depletion chains

    Science.gov (United States)

    Mosca, Pietro; Mounier, Claude

    2016-03-01

    The automatic construction of evolution chains recently implemented in GALILEE system is based on the analysis of several ENDF files : the multigroup production cross sections present in the GENDF files processed by NJOY from the ENDF evaluation, the decay file and the fission product yields (FPY) file. In this context, this paper highlights the importance of the nucleus identification to properly interconnect the data mentioned above. The first part of the paper describes the present status of the nucleus identification among the several ENDF files focusing, in particular, on the use of the excited state number and of the isomeric state number. The second part reviews the problems encountered during the automatic construction of the depletion chains using recent ENDF data. The processing of the JEFF-3.1.1, ENDF/B-VII.0 (decay and FPY) and the JEFF-3.2 (production cross section) points out problems about the compliance or not of the nucleus identifiers with the ENDF-6 format and sometimes the inconsistencies among the various ENDF files. In addition, the analysis of EAF-2003 and EAF-2010 shows some incoherence between the ZA product identifier and the reaction identifier MT for the reactions (n, pα) and (n, 2np). As a main result of this work, our suggestion is to change the ENDF format using systematically the isomeric state number to identify the nuclei. This proposal is already compliant to a huge amount ENDF data that are not in agreement with the present ENDF format. This choice is the most convenient because, ultimately, it allows one to give human readable names to the nuclei of the depletion chains.

  4. Eco-innovation Strategies in the Construction Sector: Impacts on Nanotech Innovation in the Window Chain

    Science.gov (United States)

    Andersen, M. M.; Molin, M.

    In this paper we examine the strategic response of the construction companies to the climate agenda and the emerging nanotechnologies. A case is brought on the Danish window industry. The construction industry has for a long time been considered little innovative and this also goes for the window section. It seems that a combination of an intensified focus on climate issues and the potential of nanotechnology is invoking a new innovation potential and pressure in the window industry. Specifically we identify a strategic shift among the major players towards more systemic innovations that places the window as a part of the wider energy system of the house, a trend that isn’t driven by but could be reinforced by the new nanotech opportunities that so far only play a limited strategic role.

  5. Drug Development Value Chain Constructed by Collaboration Between The SOSHO Project and The NPO BIOGRID

    Science.gov (United States)

    Inoue, Tsuyoshi; Kado, Yuji; Tokuoka, Keiji; Matsumura, Hiroyoshi; Kai, Yasushi; Mori, Yusuke; Adachi, Hiroaki; Takano, Kazufumi; Murakami, Satoshi; Fukunishi, Yoshifumi; Nakamura, Haruki; Kinoshita, Takayoshi; Nakanishi, Isao; Okuno, Yasushi; Minakata, Seiji; Sakata, Tsuneaki

    2007-03-01

    We recently established a drug development value chain collaborated by The SOSHO project (http://www.sosho.jp) and The BioGrid Project (http://www.biogrid.jp/) to accelerate new drug development. The SOSHO project provides new crystal growth methods including handling of protein crystals, and The BioGrid Project their developing software necessary for the in silico screening of promising drugs and the simulation of biological responses to proteins. We selected the two target enzymes; human hematopoietic prostaglandin D synthase (H-PGDS) and orotidine 5'-monophosphate decarboxylase from human malaria parasite plasmodium falciparum (PfOMPDC). The optimizing of HQL-79, the inhibitor for human H-PGDS and the screening of a lead compound for PfOMPDC by using in silico method are in study.

  6. Fv1 restriction and retrovirus vaccine immunity in Apobec3-deficient 129P2 mice.

    Directory of Open Access Journals (Sweden)

    Kalani Halemano

    Full Text Available Understanding the host genetics of the immune response in retrovirus infection models could provide insights for basic HIV vaccine discovery. In Friend retrovirus (FV infection of mice, Fv1 differentially inhibits N-tropic versus B-tropic FV infection by mediating a capsid-dependent post-entry block, Fv2 susceptibility governs splenomegaly induction, and Rfv3 resistance primes a stronger neutralizing antibody response due to more potent Apobec3 activity. Apobec3 polymorphisms in inbred mouse strains correlate with Rfv3 resistance and susceptibility, with one unresolved exception. The 129/OlaHsd (129P2 mouse strain is Fv2 and Rfv3 susceptible based on genotyping, but infection of 129P2 mice with B-tropic FV resulted in strong neutralizing antibody responses and no splenomegaly. Here we confirm that 129P2 mice are Fv1(nr/nr, explaining its resistance to B-tropic FV. Infection of 129P2 mice with NB-tropic FV, which can efficiently infect mice independent of Fv1 genotype, resulted in severe splenomegaly, high levels of viremia and weak neutralizing antibody responses regardless of Apobec3 status. Notably, high-dose B-tropic FV infection of 129P2 Apobec3-deficient mice induced significant adaptive immune responses and conferred high levels of protection following challenge with pathogenic NB-tropic FV. This immunological protection complemented previous studies that N-tropic FV can act as a live-attenuated vaccine in Fv1 (b/b mice. Altogether, the results obtained in 129P2 mice strengthen the conclusion that Rfv3 is encoded by Apobec3, and highlight Fv1 incompatibility as a retroviral vaccine paradigm in mice. Due to its susceptibility to disease that allows for pathogenic challenge studies, B-tropic FV infection of 129P2 mice may be a useful model to study the immunological pathways induced by retroviral capsid restriction.

  7. Spherical core-shell magnetic particles constructed by main-chain palladium N-heterocyclic carbenes

    Science.gov (United States)

    Zhao, Huaixia; Li, Liuyi; Wang, Jinyun; Wang, Ruihu

    2015-02-01

    The encapsulation of the functional species on magnetic core is a facile approach for the synthesis of core-shell magnetic materials, and surface encapsulating matrices play crucial roles in regulating their properties and applications. In this study, two core-shell palladium N-heterocyclic carbene (NHC) particles (Fe3O4@PNP1 and Fe3O4@PNP2) were prepared by a one-pot reaction of semi-rigid tripodal imidazolium salts and palladium acetate in the presence of magnetite nanoparticles. The magnetite nanoparticles are encapsulated inside the main-chain palladium, which act as cores. The conjugated effects of triphenyltriazine and triphenylbenzene in the imidazolium salts have important influence on their physical properties and catalytic performances. Fe3O4@PNP2 shows better recyclability than Fe3O4@PNP1. Unexpectedly, Pd(ii) is well maintained after six consecutive catalytic runs in Fe3O4@PNP2, and Pd(0) and Pd(ii) coexist in Fe3O4@PNP1 under the same conditions; moreover, the morphologies of these spherical core-shell particles show no significant variation after six consecutive catalytic runs.The encapsulation of the functional species on magnetic core is a facile approach for the synthesis of core-shell magnetic materials, and surface encapsulating matrices play crucial roles in regulating their properties and applications. In this study, two core-shell palladium N-heterocyclic carbene (NHC) particles (Fe3O4@PNP1 and Fe3O4@PNP2) were prepared by a one-pot reaction of semi-rigid tripodal imidazolium salts and palladium acetate in the presence of magnetite nanoparticles. The magnetite nanoparticles are encapsulated inside the main-chain palladium, which act as cores. The conjugated effects of triphenyltriazine and triphenylbenzene in the imidazolium salts have important influence on their physical properties and catalytic performances. Fe3O4@PNP2 shows better recyclability than Fe3O4@PNP1. Unexpectedly, Pd(ii) is well maintained after six consecutive catalytic runs in

  8. Antiperiodic XXZ chains with arbitrary spins: Complete eigenstate construction by functional equations in separation of variables

    CERN Document Server

    Niccoli, G

    2014-01-01

    Generic inhomogeneous integrable XXZ chains with arbitrary spins are studied by means of the quantum separation of variables (SOV) method. Within this framework, a complete description of the spectrum (eigenvalues and eigenstates) of the antiperiodic transfer matrix is derived in terms of discrete systems of equations involving the inhomogeneity parameters of the model. We show here that one can reformulate this discrete SOV characterization of the spectrum in terms of functional T-Q equations of Baxter's type, hence proving the completeness of the solutions to the associated systems of Bethe-type equations. More precisely, we consider here two such reformulations. The first one is given in terms of Q-solutions, in the form of trigonometric polynomials of a given degree $N_s$, of a one-parameter family of T-Q functional equations with an extra inhomogeneous term. The second one is given in terms of Q-solutions, again in the form of trigonometric polynomials of degree $N_s$ but with double period, of Baxter's ...

  9. Extent and Effect of Horizontal Supply Chain Collaboration among Construction SME

    Directory of Open Access Journals (Sweden)

    Liv Torjussen

    2012-01-01

    Full Text Available The majority of companies involved in value delivery in the Swedish housing industry are Small- and Medium-sized Enterprises (SME. An SME is often managed in an informal way with focus on sales and production. Many SME are also financially vulnerable as they are strongly dependent on a few key customers and key products. As variation will always exist, SME should learn to deal with variation instead of try eliminating it. This paper hypothesises that structural flexibility in SME supply chains through horizontal collaboration leads to a regional environment of economical growth from which all active SME will benefit. The hypothesis is examined through two case studies; a Swedish supplier network that has worked together six years and a four year old Norwegian supplier network. A benefit of collaboration is knowledge sharing that lessens the economical strain of keeping up with the “latest”. Other examples of collaboration are shared production resources in case of low capacity. Collaboration within supplier tier networks is considered to mark the emergence of a “collective strength” that improves individual suppliers bargaining position towards their customers. This evolution is considered an indication of the emergence of a “Lean Enterprise” within the house building sector.

  10. Antiepidermal growth factor variant III scFv fragment: effect of radioiodination method on tumor targeting and normal tissue clearance

    Energy Technology Data Exchange (ETDEWEB)

    Shankar, Sriram [Department of Radiology, Duke University Medical Center, Durham, NC 27710 (United States); Vaidyanathan, Ganesan [Department of Radiology, Duke University Medical Center, Durham, NC 27710 (United States); Kuan, C.-T. [Department of Pathology, Duke University Medical Center, Durham, NC 27710 (United States); Bigner, Darell D. [Department of Pathology, Duke University Medical Center, Durham, NC 27710 (United States); Zalutsky, Michael R. [Department of Radiology, Duke University Medical Center, Durham, NC 27710 (United States) and Department of Pathology, Duke University Medical Center, Durham, NC 27710 (United States) and Department of Biomedical Engineering, Duke University, Durham, NC 27708 (United States)]. E-mail: zalut001@duke.edu

    2006-01-15

    Introduction: MR1-1 is a single-chain Fv (scFv) fragment that binds with high affinity to epidermal growth factor receptor variant III, which is overexpressed on gliomas and other tumors but is not present on normal tissues. The objective of this study was to evaluate four different methods for labeling MR1-1 scFv that had been previously investigated for the radioiodinating of an intact anti-epidermal growth factor receptor variant III (anti-EGFRvIII) monoclonal antibody (mAb) L8A4. Methods: The MR1-1 scFv was labeled with {sup 125}I/{sup 131}I using the Iodogen method, and was also radiohalogenated with acylation agents bearing substituents that were positively charged-N-succinimidyl-3-[*I]iodo-5-pyridine carboxylate and N-succinimidyl-4-guanidinomethyl-3-[*I]iodobenzoate ([*I]SGMIB)-and negatively charged-N-succinimidyl-3-[*I]iodo-4-phosphonomethylbenzoate ([*I]SIPMB). In vitro internalization assays were performed with the U87MG{delta}EGFR cell line, and the tissue distribution of the radioiodinated scFv fragments was evaluated in athymic mice bearing subcutaneous U87MG{delta}EGFR xenografts. Results and Conclusion: As seen previously with the anti-EGFRvIII IgG mAb, retention of radioiodine activity in U87MG{delta}EGFR cells in the internalization assay was labeling method dependent, with SGMIB and SIPMB yielding the most prolonged retention. However, unlike the case with the intact mAb, the results of the internalization assays were not predictive of in vivo tumor localization capacity of the labeled scFv. Renal activity was dependent on the nature of the labeling method. With MR1-1 labeled using SIPMB, kidney uptake was highest and most prolonged; catabolism studies indicated that this uptake primarily was in the form of {epsilon}-N-3-[*I]iodo-4-phosphonomethylbenzoyl lysine.

  11. Light quality regulates flowering in FvFT1/FvTFL1 dependent manner in the woodland strawberry Fragaria vesca.

    Science.gov (United States)

    Rantanen, Marja; Kurokura, Takeshi; Mouhu, Katriina; Pinho, Paulo; Tetri, Eino; Halonen, Liisa; Palonen, Pauliina; Elomaa, Paula; Hytönen, Timo

    2014-01-01

    Control of flowering in the perennial model, the woodland strawberry (Fragaria vesca L.), involves distinct molecular mechanisms that result in contrasting photoperiodic flowering responses and growth cycles in different accessions. The F. vesca homolog of TERMINAL FLOWER1 (FvTFL1) functions as a key floral repressor that causes short-day (SD) requirement of flowering and seasonal flowering habit in the SD strawberry. In contrast, perpetual flowering F. vesca accessions lacking functional FvTFL1 show FLOWERING LOCUS T (FvFT1)-dependent early flowering specifically under long-days (LD). We show here that the end-of-day far-red (FR) and blue (B) light activate the expression of FvFT1 and the F. vesca homolog of SUPPRESSOR OF THE OVEREXPRESSION OF CONSTANS (FvSOC1) in both SD and LD strawberries, whereas low expression levels are detected in red (R) and SD treatments. By using transgenic lines, we demonstrate that FvFT1 advances flowering under FR and B treatments compared to R and SD treatments in the LD strawberry, and that FvSOC1 is specifically needed for the B light response. In the SD strawberry, flowering responses to these light quality treatments are reversed due to up-regulation of the floral repressor FvTFL1 in parallel with FvFT1 and FvSOC1. Our data highlights the central role of FvFT1 in the light quality dependent flower induction in the LD strawberry and demonstrates that FvTFL1 reverses not only photoperiodic requirements but also light quality effects on flower induction in the SD strawberry. PMID:24966865

  12. A phage-displayed chicken single-chain antibody fused to alkaline phosphatase detects Fusarium pathogens and their presence in cereal grains

    International Nuclear Information System (INIS)

    Graphical abstract: A phage-displayed chicken scFv antibody, FvSG7, binds on the surface antigen of conidiospores and the mycelia of F. verticillioides. Its fusion with alkaline phosphatase (AP) through a 218 linker displayed a 4-fold higher affinity compared with the parent scFv antibody and efficiently detected toxigenic Fusarium pathogens in cereal grains. Highlights: ► Generation of a highly reactive scFv antibody against F. verticillioides. ► Localization of the antibody binding to the surface target of F. verticillioides. ► Expression of the antibody–alkaline phosphatase (AP) fusion linked by a 218 linker. ► The antibody–AP fusion has a higher affinity than the parental antibody. ► The antibody–AP fusion detects toxigenic Fusarium pathogens in cereal grains. -- Abstract: Fusarium and its poisonous mycotoxins are distributed worldwide and are of particular interest in agriculture and food safety. A simple analytical method to detect pathogens is essential for forecasting diseases and controlling mycotoxins. This article describes a proposed method for convenient and sensitive detection of Fusarium pathogens that uses the fusion of single-chain variable fragment (scFv) and alkaline phosphatase (AP). A highly reactive scFv antibody specific to soluble cell wall-bound proteins (SCWPs) of F. verticillioides was selected from an immunized chicken phagemid library by phage display. The antibody was verified to bind on the surface of ungerminated conidiospores and mycelia of F. verticillioides. The scFv–AP fusion was constructed, and soluble expression in bacteria was confirmed. Both the antibody properties and enzymatic activity were retained, and the antigen-binding capacity of the fusion was enhanced by the addition of a linker. Surface plasmon resonance measurements confirmed that the fusion displayed 4-fold higher affinity compared with the fusion's parental scFv antibody. Immunoblot analyses showed that the fusion had good binding capacity to

  13. A phage-displayed chicken single-chain antibody fused to alkaline phosphatase detects Fusarium pathogens and their presence in cereal grains

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Zu-Quan [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); Li, He-Ping [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); Zhang, Jing-Bo [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); Huang, Tao [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); Liu, Jin-Long; Xue, Sheng [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); Wu, Ai-Bo [Institute for Agri-food Standards and Testing Technology, Laboratory of Quality and Safety Risk Assessment for Agro-products, Ministry of Agriculture, Shanghai Academy of Agricultural Sciences, 1000 Jinqi Road, Shanghai 201403 (China); Liao, Yu-Cai, E-mail: ycliao06@yahoo.com.cn [Molecular Biotechnology Laboratory of Triticeae Crops, Huazhong Agricultural University, Wuhan 430070 (China); College of Plant Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); National Center of Plant Gene Research, Wuhan 430070 (China)

    2013-02-18

    Graphical abstract: A phage-displayed chicken scFv antibody, FvSG7, binds on the surface antigen of conidiospores and the mycelia of F. verticillioides. Its fusion with alkaline phosphatase (AP) through a 218 linker displayed a 4-fold higher affinity compared with the parent scFv antibody and efficiently detected toxigenic Fusarium pathogens in cereal grains. Highlights: ► Generation of a highly reactive scFv antibody against F. verticillioides. ► Localization of the antibody binding to the surface target of F. verticillioides. ► Expression of the antibody–alkaline phosphatase (AP) fusion linked by a 218 linker. ► The antibody–AP fusion has a higher affinity than the parental antibody. ► The antibody–AP fusion detects toxigenic Fusarium pathogens in cereal grains. -- Abstract: Fusarium and its poisonous mycotoxins are distributed worldwide and are of particular interest in agriculture and food safety. A simple analytical method to detect pathogens is essential for forecasting diseases and controlling mycotoxins. This article describes a proposed method for convenient and sensitive detection of Fusarium pathogens that uses the fusion of single-chain variable fragment (scFv) and alkaline phosphatase (AP). A highly reactive scFv antibody specific to soluble cell wall-bound proteins (SCWPs) of F. verticillioides was selected from an immunized chicken phagemid library by phage display. The antibody was verified to bind on the surface of ungerminated conidiospores and mycelia of F. verticillioides. The scFv–AP fusion was constructed, and soluble expression in bacteria was confirmed. Both the antibody properties and enzymatic activity were retained, and the antigen-binding capacity of the fusion was enhanced by the addition of a linker. Surface plasmon resonance measurements confirmed that the fusion displayed 4-fold higher affinity compared with the fusion's parental scFv antibody. Immunoblot analyses showed that the fusion had good binding

  14. Effects of interlinker sequences on the biological properties of bispecific single-chain antibodies

    Institute of Scientific and Technical Information of China (English)

    FANG Min; JIANG Xin; YANG Zhi; YIN Changcheng; LI Hua; ZHAO Rui; ZHANG Zhong; LIN Qing; HUANG Hualiang

    2003-01-01

    Single-chain bispecific antibody (scBsAb) is one of the promising genetic engineering antibody formats for clinical application. But the effects of interlinker sequences on the biological properties of bispecific single-chain antibodies have not been studied in detail. Three interlinker sequences were designed and synthesized, and denominated as Fc, HSA, 205C′, respectively. Universal vectors with these different interlinker sequences for scBsAb expression in E. coli were constructed. A model scBsAb based on a reshaped single-chain antibody (scFv) against human CD3 and a scFv directed against human ovarian carcinoma were generated and expressed in E. coli. The results of SDS-PAGE and Western blot showed that the different interlinker sequences did not affect the expression levelof scBsAb. However, as demonstrated by ELISA and pharmacokinetics studies performed in mice, scBsAbs with different interlinker sequences had difference in the antigen-binding activities and terminal half-life time (T1/2β) in vivo, the interlinker HSA could remarkably prolong the retention time of scBsAb in blood. These results indicated that the peptide sequence of interlinker could affect important biological properties of scBsAb, such as antigen-binding properties and stability in vivo. So, selection of an appropriate interlinker sequence is very important for scBsAb construction. Optimal interlinker can bring scBsAb biologicalproperties more suitable for clinical application.

  15. Generation of a Novel Bacteriophage Library Displaying scFv Antibody Fragments from the Natural Buffalo Host to Identify Antigens from Adult Schistosoma japonicum for Diagnostic Development.

    Directory of Open Access Journals (Sweden)

    Christopher G Hosking

    2015-12-01

    Full Text Available The development of effective diagnostic tools will be essential in the continuing fight to reduce schistosome infection; however, the diagnostic tests available to date are generally laborious and difficult to implement in current parasite control strategies. We generated a series of single-chain antibody Fv domain (scFv phage display libraries from the portal lymph node of field exposed water buffaloes, Bubalus bubalis, 11-12 days post challenge with Schistosoma japonicum cercariae. The selected scFv-phages showed clear enrichment towards adult schistosomes and excretory-secretory (ES proteins by immunofluorescence, ELISA and western blot analysis. The enriched libraries were used to probe a schistosome specific protein microarray resulting in the recognition of a number of proteins, five of which were specific to schistosomes, with RNA expression predominantly in the adult life-stage based on interrogation of schistosome expressed sequence tags (EST. As the libraries were enriched by panning against ES products, these antigens may be excreted or secreted into the host vasculature and hence may make good targets for a diagnostic assay. Further selection of the scFv library against infected mouse sera identified five soluble scFv clones that could selectively recognise soluble whole adult preparations (SWAP relative to an irrelevant protein control (ovalbumin. Furthermore, two of the identified scFv clones also selectively recognised SWAP proteins when spiked into naïve mouse sera. These host B-cell derived scFvs that specifically bind to schistosome protein preparations will be valuable reagents for further development of a cost effective point-of-care diagnostic test.

  16. Generation of a Novel Bacteriophage Library Displaying scFv Antibody Fragments from the Natural Buffalo Host to Identify Antigens from Adult Schistosoma japonicum for Diagnostic Development.

    Science.gov (United States)

    Hosking, Christopher G; McWilliam, Hamish E G; Driguez, Patrick; Piedrafita, David; Li, Yuesheng; McManus, Donald P; Ilag, Leodevico L; Meeusen, Els N T; Veer, Michael J de

    2015-12-01

    The development of effective diagnostic tools will be essential in the continuing fight to reduce schistosome infection; however, the diagnostic tests available to date are generally laborious and difficult to implement in current parasite control strategies. We generated a series of single-chain antibody Fv domain (scFv) phage display libraries from the portal lymph node of field exposed water buffaloes, Bubalus bubalis, 11-12 days post challenge with Schistosoma japonicum cercariae. The selected scFv-phages showed clear enrichment towards adult schistosomes and excretory-secretory (ES) proteins by immunofluorescence, ELISA and western blot analysis. The enriched libraries were used to probe a schistosome specific protein microarray resulting in the recognition of a number of proteins, five of which were specific to schistosomes, with RNA expression predominantly in the adult life-stage based on interrogation of schistosome expressed sequence tags (EST). As the libraries were enriched by panning against ES products, these antigens may be excreted or secreted into the host vasculature and hence may make good targets for a diagnostic assay. Further selection of the scFv library against infected mouse sera identified five soluble scFv clones that could selectively recognise soluble whole adult preparations (SWAP) relative to an irrelevant protein control (ovalbumin). Furthermore, two of the identified scFv clones also selectively recognised SWAP proteins when spiked into naïve mouse sera. These host B-cell derived scFvs that specifically bind to schistosome protein preparations will be valuable reagents for further development of a cost effective point-of-care diagnostic test. PMID:26684756

  17. Nanoscale superstructures assembled by polymerase chain reaction (PCR): programmable construction, structural diversity, and emerging applications.

    Science.gov (United States)

    Kuang, Hua; Ma, Wei; Xu, Liguang; Wang, Libing; Xu, Chuanlai

    2013-11-19

    Polymerase chain reaction (PCR) is an essential tool in biotechnology laboratories and is becoming increasingly important in other areas of research. Extensive data obtained over the last 12 years has shown that the combination of PCR with nanoscale dispersions can resolve issues in the preparation DNA-based materials that include both inorganic and organic nanoscale components. Unlike conventional DNA hybridization and antibody-antigen complexes, PCR provides a new, effective assembly platform that both increases the yield of DNA-based nanomaterials and allows researchers to program and control assembly with predesigned parameters including those assisted and automated by computers. As a result, this method allows researchers to optimize to the combinatorial selection of the DNA strands for their nanoparticle conjugates. We have developed a PCR approach for producing various nanoscale assemblies including organic motifs such as small molecules, macromolecules, and inorganic building blocks, such as nanorods (NRs), metal, semiconductor, and magnetic nanoparticles (NPs). We start with a nanoscale primer and then modify that building block using the automated steps of PCR-based assembly including initialization, denaturation, annealing, extension, final elongation, and final hold. The intermediate steps of denaturation, annealing, and extension are cyclic, and we use computer control so that the assembled superstructures reach their predetermined complexity. The structures assembled using a small number of PCR cycles show a lower polydispersity than similar discrete structures obtained by direct hybridization between the nanoscale building blocks. Using different building blocks, we assembled the following structural motifs by PCR: (1) discrete nanostructures (NP dimers, NP multimers including trimers, pyramids, tetramers or hexamers, etc.), (2) branched NP superstructures and heterochains, (3) NP satellite-like superstructures, (4) Y-shaped nanostructures and DNA

  18. pMINERVA: A donor-acceptor system for the in vivo recombineering of scFv into IgG molecules.

    Science.gov (United States)

    Batonick, M; Kiss, M M; Fuller, E P; Magadan, C M; Holland, E G; Zhao, Q; Wang, D; Kay, B K; Weiner, M P

    2016-04-01

    Phage display is the most widely used method for selecting binding molecules from recombinant antibody libraries. However, validation of the phage antibodies often requires early production of the cognate full-length immunoglobulin G (IgG). The conversion of phage library outputs to a full immunoglobulin via standard subcloning is time-consuming and limits the number of clones that can be evaluated. We have developed a novel system to convert scFvs from a phage display vector directly into IgGs without any in vitro subcloning steps. This new vector system, named pMINERVA, makes clever use of site-specific bacteriophage integrases that are expressed in Escherichia coli and intron splicing that occurs within mammalian cells. Using this system, a phage display vector contains both bacterial and mammalian regulatory regions that support antibody expression in E. coli and mammalian cells. A single-chain variable fragment (scFv) antibody is expressed on the surface of bacteriophage M13 as a genetic fusion to the gpIII coat protein. The scFv is converted to an IgG that can be expressed in mammalian cells by transducing a second E. coli strain. In that strain, the phiC31 recombinase fuses the heavy chain constant domain from an acceptor plasmid to the heavy chain variable domain and introduces controlling elements upstream of the light chain variable domain. Splicing in mammalian cells removes a synthetic intron containing the M13 gpIII gene to produce the fusion of the light chain variable domain to the constant domain. We show that phage displaying a scFv and recombinant IgGs generated using this system are expressed at wild-type levels and retain normal function. Use of the pMINERVA completely eliminates the labor-intensive subcloning and DNA sequence confirmation steps currently needed to convert a scFv into a functional IgG Ab. PMID:26851519

  19. THE CONSTRUCTION AND EXPRESSION OF THE MURINE SCFV GENE IN E. COLI AGAINST HUMAN CERVICAL CANCER

    Institute of Scientific and Technical Information of China (English)

    Wang Ying; Chen Wei; Li Xu

    2006-01-01

    Objective To obtain the gene of murine Single chain Fv fragment (ScFv) against human cervical cancer and to express it in E. coli. Methods The variable region gene fragments of the heavy and light chains, which were amplified respectively using recombinant DNA techniques from CsA125 hybridoma cells, were spliced together through a flexible linker to ScFv against human cervical cancer. The ScFv genes were then cloned into expression vector pCANTAB 5E and expressed in E. coli HB2151 and TG1 respectively. The soluble ScFv were characterized by SDS PAGE and Western blot. The antigen-binding activities of the soluble and phage displayed ScFv were assayed by ELISA and cell immunohistochemical analysis. Results The expressed ScFv antibodies were soluble and phage displayed. The soluble ScFv secreted and expressed in E. coli HB2151 induced by IPTG were confirmed with SDS-PAGE, Western blot and ELISA. The specific binding capacity of the soluble and phage displayed ScFv to the surface associated antigen of human cervical cancer cell line was further confirmed with immunohistochemical studies. Conclusion The soluble and phage displayed ScFv expressed in E. coli against human cervical cancer showed high, specific affinity for the cervical cancer cell line surface associated antigen.

  20. BIM技术在施工项目供应链管理中的应用%Application of BIM in Supply Chain Management of Construction Projects

    Institute of Scientific and Technical Information of China (English)

    俞启元; 吕玉惠; 张尚

    2016-01-01

    将供应链管理、系统集成和信息技术交叉起来,研究BIM技术在施工项目供应链管理中的应用。首先,阐述施工项目供应链管理的信息传递方式及BIM技术的特点;然后,基于对信息传递的认识,提出将BIM技术应用于施工项目供应链管理的基本路径;最后,设计基于BIM技术的施工项目供应链管理信息系统基本架构。%This paper integrates the supply chain management,system integration and information technology, studies the BIM application in supply chain management of construction project. Firstly,expounds the information transmission mode in the supply chain management of construction project and the characteristics of BIM. Secondly,based on cognition of information transfer,puts forward the basic path of applying BIM in supply chain management of construction project. Finally,designs the basic structure of supply chain management information system of construction project based on BIM Technology.

  1. Production and characterization of a single-chain antibody of anti-CD3%抗CD3单链抗体基因克隆表达及生物活性鉴定

    Institute of Scientific and Technical Information of China (English)

    陈秀芹; 阎锡蕴

    2003-01-01

    The genes encoding antibody heavy and light chain variable regions(VH and VL)were cloned by RT-PCR from OKT3 hybridoma cells,which produced anti-CD3 moleclonal antibody.The VH and VL genes were fused and become a single chain Fv(scFv).The scFv gene was cloned into pCANTAB5E vector and expressed on bacterial phage surface.By three panning rounds,we have obtained two single-chain antibodys that specific for CD3.The antiCD3 scFv wil be a reagent fox diagnosis and therapy of immuno-disorder.

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

    Science.gov (United States)

    Cheung, Nai-Kong; Guo, Hong-Fen

    2010-02-23

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

  3. Evaluation of anti-HER2 scFv-conjugated PLGA-PEG nanoparticles on 3D tumor spheroids of BT474 and HCT116 cancer cells

    Science.gov (United States)

    Thuy Duong Le, Thi; Pham, Thu Hong; Nghia Nguyen, Trong; Giang Ngo, Thi Hong; Nhung Hoang, Thi My; Huan Le, Quang

    2016-06-01

    Three-dimensional culture cells (spheroids) are one of the multicellular culture models that can be applied to anticancer chemotherapeutic development. Multicellular spheroids more closely mimic in vivo tumor-like patterns of physiologic environment and morphology. In previous research, we designed docetaxel-loaded pegylated poly(D, L-lactide-co-glycolide) nanoparticles conjugated with anti-HER2 single chain antibodies (scFv-Doc-PLGA-PEG) and evaluated them in 2D cell culture. In this study, we continuously evaluate the cellular uptake and cytotoxic effect of scFv-Doc-PLGA-PEG on a 3D tumor spheroid model of BT474 (HER2-overexpressing) and HCT116 (HER2-underexpressing) cancer cells. The results showed that the nanoparticle formulation conjugated with scFv had a significant internalization effect on the spheroids of HER2-overexpressing cancer cells as compared to the spheroids of HER2-underexpressing cancer cells. Therefore, cytotoxic effects of targeted nanoparticles decreased the size and increased necrotic score of HER2-overexpressing tumor spheroids. Thus, these scFv-Doc-PLGA-PEG nanoparticles have potential for active targeting for HER2-overexpressing cancer therapy. In addition, BT474 and HCT116 spheroids can be used as a tumor model for evaluation of targeting therapies.

  4. Preparation and diagnostic use of a novel recombinant single-chain antibody against rabies virus glycoprotein.

    Science.gov (United States)

    Yuan, Ruosen; Chen, Xiaoxu; Chen, Yan; Gu, Tiejun; Xi, Hualong; Duan, Ye; Sun, Bo; Yu, Xianghui; Jiang, Chunlai; Liu, Xintao; Wu, Chunlai; Kong, Wei; Wu, Yongge

    2014-02-01

    Rabies virus (RABV) causes a fatal infectious disease, but effective protection may be achieved with the use of rabies immunoglobulin and a rabies vaccine. Virus-neutralizing antibodies (VNA), which play an important role in the prevention of rabies, are commonly evaluated by the RABV neutralizing test. For determining serum VNA levels or virus titers during the RABV vaccine manufacturing process, reliability of the assay method is highly important and mainly dependent on the diagnostic antibody. Most diagnostic antibodies are monoclonal antibodies (mAbs) made from hybridoma cell lines and are costly and time consuming to prepare. Thus, production of a cost-effective mAb for determining rabies VNA levels or RABV titers is needed. In this report, we describe the prokaryotic production of a RABV-specific single-chain variable fragment (scFv) protein with a His-tag (scFv98H) from a previously constructed plasmid in a bioreactor, including the purification and refolding process as well as the functional testing of the protein. The antigen-specific binding characteristics, affinity, and relative affinity of the purified protein were tested. The scFv98H antibody was compared with a commercial RABV nucleoprotein mAb for assaying the VNA level of anti-rabies serum samples from different sources or testing the growth kinetics of RABV strains for vaccine manufactured in China. The results indicated that scFv98H may be used as a novel diagnostic tool to assay VNA levels or virus titers and may be used as an alternative for the diagnostic antibody presently employed for these purposes. PMID:24241896

  5. A phage-displayed chicken single-chain antibody fused to alkaline phosphatase detects Fusarium pathogens and their presence in cereal grains.

    Science.gov (United States)

    Hu, Zu-Quan; Li, He-Ping; Zhang, Jing-Bo; Huang, Tao; Liu, Jin-Long; Xue, Sheng; Wu, Ai-Bo; Liao, Yu-Cai

    2013-02-18

    Fusarium and its poisonous mycotoxins are distributed worldwide and are of particular interest in agriculture and food safety. A simple analytical method to detect pathogens is essential for forecasting diseases and controlling mycotoxins. This article describes a proposed method for convenient and sensitive detection of Fusarium pathogens that uses the fusion of single-chain variable fragment (scFv) and alkaline phosphatase (AP). A highly reactive scFv antibody specific to soluble cell wall-bound proteins (SCWPs) of F. verticillioides was selected from an immunized chicken phagemid library by phage display. The antibody was verified to bind on the surface of ungerminated conidiospores and mycelia of F. verticillioides. The scFv-AP fusion was constructed, and soluble expression in bacteria was confirmed. Both the antibody properties and enzymatic activity were retained, and the antigen-binding capacity of the fusion was enhanced by the addition of a linker. Surface plasmon resonance measurements confirmed that the fusion displayed 4-fold higher affinity compared with the fusion's parental scFv antibody. Immunoblot analyses showed that the fusion had good binding capacity to the components from SCWPs of F. verticillioides, and enzyme-linked immunosorbent assays revealed that the detection limit of the fungus was below 10(-2) μg mL(-1), superior to the scFv antibody. The fusion protein was able to detect fungal concentrations as low as 10(-3) mg g(-1) of maize grains in both naturally and artificially contaminated samples. Thus, the fusion can be applied in rapid and simple diagnosis of Fusarium contamination in field and stored grain or in food. PMID:23374219

  6. 应急物流供应链构建研究%Research on the Construction of Emergency Logistics Supply Chain

    Institute of Scientific and Technical Information of China (English)

    刘玲丽

    2014-01-01

    文中在阐释应急物流及其供应链概念的基础上,分析了应急物流供应链的特点及流程,构建了应急物流供应链结构模型,探讨了应急物流供应链的构建原则、参与主体构成、协同机制的建立及信息平台的构建等相关问题。%Base on the definition of emergency logistics supply chain ,this paper analyses it's characteristic and flow.The paper also structures a model of emergency logistics supply chain and discusses the relevant questions about the emergency logistics supply chain construction principles ,it's main participants ,coordination mechanism ,information platform construction and so on.

  7. Proceedings of the Canadian Institute conference on supply chain management in the oil and gas industry : major capital construction projects, maintenance, repair and operations

    International Nuclear Information System (INIS)

    Many companies are now being forced to focus on careful budgeting to ensure that the capital costs of large-scale construction projects do not exceed their budgets. Operators are now investigating the role of supply chain management in reducing project costs. This conference provided a forum for the discussion of issues related to large construction projects for supply chain management in the oil and gas industry. Participants at the conference discussed methods of negotiating with contractors in order to manage higher prices for steel and other commodities. Best practices for maintaining effective purchaser-contractor relations were discussed along with cost benchmarks in contracts and management planning techniques for supply chain processes. The benefits of adopting vendor-managed inventory systems were also examined. Sourcing strategies were presented and issues related to transportation were reviewed along with various planning models. The conference featured 16 presentations. tabs., figs

  8. Potential therapeutic strategy for non-Hodgkin lymphoma by anti-CD20scFvFc/CD28/CD3zeta gene tranfected T cells

    Directory of Open Access Journals (Sweden)

    Zheng Yihu

    2010-09-01

    Full Text Available Abstract Background Anti-CD20 monoclonal antibody treatment has not only increased survival and cure rates in many non-Hodgkin lymphomas, but also has prompted an explosion in the development of novel antibodies and biologically active substances with specific cellular targets in the field of malignancies treatment. Since the robust immune responses are elicited by the gene-modified T cells, gene based T cell therapy may also provide a powerful tool for cancer immunotherapy. Methods In this study, we developed a vector construction encoding a chimeric T cell receptor that recognizes the CD20 antigen and delivers co-stimulatory signals to achieve T cell activation. One non-Hodgkin lymphoma cell line Raji cells co-cultured with peripheral blood-derived T cells were stably transfected with anti-CD20scFvFc/CD28/CD3zeta gene or anti-CD20scFvFc gene. T cells expressing anti-CD20scFvFc/CD28/CD3zeta or anti-CD20scFvFc gene co-cultured with CD20 positive Raji cells for different times. Cell lysis assay was carried by [3H]TdR release assay. The expressions of Fas, Bcl-2 and Caspase-3 of Raji cells were detected by flow cytometric. The secretion of IFN-gamma and IL-2 in co-culture medium was tested by ELISA assay. Activity of AP-1 was analyzed by EMSA. Results Following efficient transduction of peripheral blood-derived T cells with anti-CD20scFvFc/CD28/CD3zeta gene, an obvious cell lysis of Raji cells was observed in co-culture. T cells transduced anti-CD20scFvFc/CD28/CD3zeta gene had superior secretion of IFN-gamma and IL-2 compared to T cells transduced anti-CD20scFvFc gene. Also it led to a much stronger Fas-induced apoptosis signaling transduction in target cancer cells. Conclusion So adoptively T cells transduced anti-CD20scFvFc/CD28/CD3zeta gene mediates enhanced anti-tumor activities against CD20 positive tumor cells, suggesting a potential of gene-based immunotherapy for non-Hodgkin lymphoma.

  9. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2008-07-01

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magnetic activated cell sorting (MACS) and fluorescence activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks resulting in a panel of scFvs specific for the target antigen.

  10. Construction and Screening of Antigen Targeted Immune Yeast Surface Display Antibody Libraries

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Keith D.; Pefaur, Noah B.; Baird, Cheryl L.

    2009-08-02

    These protocols describe a yeast surface display-based process for the rapid selection of antibodies from immunized mice, eliminating the need for creating and screening hybridoma fusions. A yeast surface display library of single-chain antibody fragments (scFvs) is created from antigen-binding B cells from the splenocytes of immunized mice. The antigen targeted library is then screened for antigen specific scFv by magneticactivated cell sorting (MACS) and fluorescence-activated cell sorting (FACS). Library construction and screening can be accomplished in as little as 2 weeks, resulting in a panel of scFvs specific for the target antigen.

  11. Toward low-cost affinity reagents: lyophilized yeast-scFv probes specific for pathogen antigens.

    Directory of Open Access Journals (Sweden)

    Sean A Gray

    Full Text Available The generation of affinity reagents, usually monoclonal antibodies, remains a critical bottleneck in biomedical research and diagnostic test development. Recombinant antibody-like proteins such as scFv have yet to replace traditional monoclonal antibodies in antigen detection applications, in large part because of poor performance of scFv in solution. To address this limitation, we have developed assays that use whole yeast cells expressing scFv on their surfaces (yeast-scFv in place of soluble purified scFv or traditional monoclonal antibodies. In this study, a nonimmune library of human scFv displayed on the surfaces of yeast cells was screened for clones that bind to recombinant cyst proteins of Entamoeba histolytica, an enteric pathogen of humans. Selected yeast-scFv clones were stabilized by lyophilization and used in detection assay formats in which the yeast-scFv served as solid support-bound monoclonal antibodies. Specific binding of antigen to the yeast-scFv was detected by staining with rabbit polyclonal antibodies. In flow cytometry-based assays, lyophilized yeast-scFv reagents retained full binding activity and specificity for their cognate antigens after 4 weeks of storage at room temperature in the absence of desiccants or stabilizers. Because flow cytometry is not available to all potential assay users, an immunofluorescence assay was also developed that detects antigen with similar sensitivity and specificity. Antigen-specific whole-cell yeast-scFv reagents can be selected from nonimmune libraries in 2-3 weeks, produced in vast quantities, and packaged in lyophilized form for extended shelf life. Lyophilized yeast-scFv show promise as low cost, renewable alternatives to monoclonal antibodies for diagnosis and research.

  12. Isolation and characterisation of a human-like antibody fragment (scFv) that inactivates VEEV in vitro and in vivo.

    Science.gov (United States)

    Rülker, Torsten; Voß, Luzie; Thullier, Philippe; O' Brien, Lyn M; Pelat, Thibaut; Perkins, Stuart D; Langermann, Claudia; Schirrmann, Thomas; Dübel, Stefan; Marschall, Hans-Jürgen; Hust, Michael; Hülseweh, Birgit

    2012-01-01

    Venezuelan equine encephalitis virus (VEEV) belongs to the Alphavirus genus and several species of this family are pathogenic to humans. The viruses are classified as potential agents of biological warfare and terrorism and sensitive detection as well as effective prophylaxis and antiviral therapies are required.In this work, we describe the isolation of the anti-VEEV single chain Fragment variable (scFv), ToR67-3B4, from a non-human primate (NHP) antibody gene library. We report its recloning into the bivalent scFv-Fc format and further immunological and biochemical characterisation.The scFv-Fc ToR67-3B4 recognised viable as well as formalin and ß-propionolactone (ß-Pl) inactivated virus particles and could be applied for immunoblot analysis of VEEV proteins and immuno-histochemistry of VEEV infected cells. It detected specifically the viral E1 envelope protein of VEEV but did not react with reduced viral glycoprotein preparations suggesting that recognition depends upon conformational epitopes. The recombinant antibody was able to detect multiple VEEV subtypes and displayed only marginal cross-reactivity to other Alphavirus species except for EEEV. In addition, the scFv-Fc fusion described here might be of therapeutic use since it successfully inactivated VEEV in a murine disease model. When the recombinant antibody was administered 6 hours post challenge, 80% to 100% of mice survived lethal VEEV IA/B or IE infection. Forty to sixty percent of mice survived when scFv-Fc ToR67-3B4 was applied 6 hours post challenge with VEEV subtypes II and former IIIA. In combination with E2-neutralising antibodies the NHP antibody isolated here could significantly improve passive protection as well as generic therapy of VEE. PMID:22666347

  13. A conjugate of an anti-midkine single-chain variable fragment to doxorubicin inhibits tumor growth

    International Nuclear Information System (INIS)

    Doxorubicin (DOX) was conjugated to a single-chain variable fragment (scFv) against human midkine (MK), and the conjugate (scFv-DOX) was used to target the chemotherapeutic agent to a mouse solid tumor model in which the tumor cells expressed high levels of human MK. The His-tagged recombinant scFv was expressed in bacteria, purified by metal affinity chromatography, and then conjugated to DOX using oxidative dextran (Dex) as a linker. The molecular formula of this immunoconjugate was scFv(Dex)1.3(DOX)20. In vitro apoptosis assays showed that the scFv-DOX conjugate was more cytotoxic against MK-transfected human adenocarcinoma cells (BGC823-MK) than untransfected cells (55.3 ± 2.4 vs 22.4 ± 3.8%) for three independent experiments. Nude mice bearing BGC823-MK solid tumors received scFv-DOX or equivalent doses of scFv + DOX for 2 weeks and tumor growth was more effectively inhibited by the scFv-DOX conjugate than by scFv + DOX (51.83% inhibition vs 40.81%). Histological analysis of the tumor tissues revealed that the highest levels of DOX accumulated in tumors from mice treated with scFv-DOX and this resulted in more extensive tumor cell death than in animals treated with the equivalent dose of scFv + DOX. These results show that the scFv-DOX conjugate effectively inhibited tumor growth in vivo and suggest that antigen-specific scFv may be competent drug-carriers

  14. Detection and characterization of cry1Ac transgene construct in Bt cotton: multiple polymerase chain reaction approach.

    Science.gov (United States)

    Singh, Chandra K; Ojha, Abhishek; Kachru, Devendra N

    2007-01-01

    To comply with international labeling regulations for genetically modified (GM) crops and food, and to enable proper identification of GM organisms (GMOs), effective methodologies and reliable approaches are needed. The spurious and unapproved GM planting has contributed to crop failures and commercial losses. To ensure effective and genuine GM cultivation, a methodology is needed to detect and identify the trait of interest and concurrently evaluate the structural and functional stability of the transgene insert. A multiple polymerase chain reaction (PCR) approach was developed for detection, identification, and gene stability confirmation of cry1Ac transgene construct in Bt cotton. As many as 9 samples of Bt cotton hybrid seeds comprising 3 approved Bt hybrids, MECH-12Bt, MECH-162Bt, MECH-184Bt, and a batch of 6 nonapproved Bt hybrids were tested. Initially, single standard PCR assays were run to amplify predominant GM DNA sequences (CaMV 35S promoter, nos terminator, and npt-II marker gene); a housekeeping gene, Gossypium hirsutum fiber-specific acyl carrier protein gene (acp1); a trait-specific transgene (cry1Ac); and a sequence of 7S 3' transcription terminator which specifically borders with 3' region of cry1Ac transgene cassette. The concurrent amplification of all sequences of the entire cassette was performed by 3 assays, duplex, triplex, and quadruplex multiplex PCR assays, under common assay conditions. The identity of amplicons was reconfirmed by restriction endonuclease digestion profile. The 2 distinct transgene cassettes, cry1Ac and npt-II, of the Bt cotton were amplified using the respective forward primer of promoter and reverse primer of terminator. The resultant amplicons were excised, eluted, and purified. The purified amplicons served as template for nested PCR assays. The nested PCR runs confirmed the transgene construct orientation and identity. The limit of detection as established by our assay for GM trait (cry1Ac) was 0.1%. This approach

  15. Reflection on Construction Project's Implementation of Supply Chain Management Model%建筑工程实施供应链管理模式的思考

    Institute of Scientific and Technical Information of China (English)

    刘柄延

    2015-01-01

    Transformed from the old management model which used to be implemented in China's previous construction engineering, the found of supply chain management model is beneficial for the establishment of new enterprise manage⁃ment model in China’s construction enterprises. The article has studied the main idea of comprehensive supply chain management concept, on this basis it puts forward some advices for the practical work in the concrete implementation of supply chain management model in projects, and conducts certain analysis of how to implement supply chain management in detail in engineering projects, hoping to provide some reference for building industry in the establishment of supply chain integration framework and the construction of supply chain management model.%供应链管理模式是对中国过去的建筑工程实施的管理模式的变革,有利于促进我国建筑企业新型企业管理模式的构建。本文研究了综合供应链管理概念的中心思想,对项目中具体实施供应链管理模式的工作提出了一些建议,对工程项目详细实施供应链管理做了一定的分析,以期为建筑业建立供应链整合架构和供应链管理模式提供一定的参考。

  16. 绿色供应链进程下建筑材料管理探究%Study on Construction Materials Management in Green Supply Chain Process

    Institute of Scientific and Technical Information of China (English)

    段文凤; 董金辉

    2015-01-01

    Through tracing the origin of green supply chain, this paper analyzed the current situation of material management in construction, discussed the problems of procuring and using building materials in the green construction, recycling the construction waste in green scrap under the green supply chain process in construction industry, proposed the mode to play the initiative of the green supply chain members from the perspective of recycled building materials manufacturers, material suppliers and engineering contractors.%本文通过追溯绿色供应链的起源,并对建筑材料管理现状进行分析,论述了建设行业绿色供应链进程下建筑材料在绿色施工中的采购使用问题和绿色报废中的建筑材料回收问题,并分别从再生建材制造商、材料供应商、工程承包商的角度提出了发挥绿色供应链各成员能动性的途径。

  17. Light quality regulates flowering in FvFT1/FvTFL1 dependent manner in the woodland strawberry Fragaria vesca

    OpenAIRE

    Rantanen, Marja; Kurokura, Takeshi; Mouhu, Katriina; Pinho, Paulo; Tetri, Eino; Halonen, Liisa; Palonen, Pauliina; Elomaa, Paula; Hytönen, Timo

    2014-01-01

    Control of flowering in the perennial model, the woodland strawberry (Fragaria vesca L.), involves distinct molecular mechanisms that result in contrasting photoperiodic flowering responses and growth cycles in different accessions. The F. vesca homolog of TERMINAL FLOWER1 (FvTFL1) functions as a key floral repressor that causes short-day (SD) requirement of flowering and seasonal flowering habit in the SD strawberry. In contrast, perpetual flowering F. vesca accessions lacking functional FvT...

  18. FvSO regulates vegetative hyphal fusion, asexual growth, fumonisin B1 production, and virulence in Fusarium verticillioides.

    Science.gov (United States)

    Guo, Li; Wenner, Nancy; Kuldau, Gretchen A

    2015-12-01

    Hyphal anastomosis is a hallmark of filamentous fungi and plays vital roles including cellular homoeostasis, interhyphal communication and nutrient translocation. Here we identify a gene, FvSO, in Fusarium verticillioides, a filamentous ascomycete causing maize ear and stalk rot and producing fumonisin mycotoxins. FvSO, like its Neurospora crassa homologue SO, is required for vegetative hyphal fusion. It is also essential for normal vegetative growth, sporulation, and pathogenesis. FvSO encodes a predicted WW domain protein and shares 70 % protein sequence identity with N. crassa SO. FvSO deletion mutants (ΔFvSO) had abnormal distribution of conidia size, and conidia of ΔFvSO germinated much later and slower than wild type. ΔFvSO was deficient in hyphal anastomosis, had slower radial growth and produced less fungal biomass than wild type. ΔFvSO were unable to perform anastomosis, a key feature of filamentous fungi. Interestingly, production of fumonisin B1 by ΔFvSO was significantly reduced compared to wild type. Additionally, ΔFvSO was nonpathogenic to corn ears, stalks and seedlings, likely due to defective growth and development. In conclusion, FvSO is essential for vegetative hyphal fusion and is required for normal vegetative growth and sporulation, normal levels of fumonisin production and pathogenicity in F. verticillioides. The pleiotropic nature of ΔFvSO phenotypes suggests that FvSO is likely involved in certain signalling pathways that regulate multiple cellular functions. PMID:26615739

  19. Analysis on construction of the Supply Chain and Brand of Guangxi Characteristic Economic Agricultural Products——A Case Study of Guilin,China

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Firstly,it is briefly introduced that Guangxi characteristic economic agricultural products are currently characterized by typical regional features and strong competitiveness.Then,Guilin is taken as an example,the problems existing in constructing the supply chain and brand of Guilin characteristic economic agricultural products are pointed out after the analysis of its current situations,the problems include weak agricultural production infrastructure,weak deep products processing capacity,low modernization level of storage and transportation as well as short of fixed marketing channels;meanwhile,brand construction faces the problems of little top brands and few differentiated products.Finally,some measures are proposed to solve these problems from the perspectives of establishing the management idea of agricultural products supply chain and enhancing the awareness to protect agricultural products brand,it is also suggested that these measures be extended to the whole Guangxi autonomous region in order to explore the way of the development of Guangxi characteristic agriculture.

  20. Fusion protein of single-chain variable domain fragments for treatment of myasthenia gravis

    Institute of Scientific and Technical Information of China (English)

    Fangfang Li; Fanping Meng; Quanxin Jin; Changyuan Sun; Yingxin Li; Honghua Li; Songzhu Jin

    2014-01-01

    Single-chain variable domain fragment (scFv) 637 is an antigen-specific scFv of myasthenia gravis. In this study, scFv and human serum albumin genes were conjugated and the fusion pro-tein was expressed in Pichia pastoris. The afifnity of scFv-human serum albumin fusion protein to bind to acetylcholine receptor at the neuromuscular junction of human intercostal muscles was detected by immunolfuorescence staining. The ability of the fusion protein to block myas-thenia gravis patient sera binding to acetylcholine receptors and its stability in healthy serum were measured by competitive ELISA. The results showed that the inhibition rate was 2.0-77.4%, and the stability of fusion protein in static healthy sera was about 3 days. This approach suggests the scFv-human serum albumin is a potential candidate for speciifc immunosuppressive therapy of myasthenia gravis.

  1. McKinsey 7S Model for Supply Chain Management of Local SMEs Construction Business in Upper Northeast Region of Thailand

    OpenAIRE

    Thanaphan Naipinit; Somkier Kojchavivong; Vorawit Kowittayakorn; Thongphon Promsaka Na Sakolnakorn

    2014-01-01

    The purpose of this study is to study the successful business strategies and the guidelines for the management strategies of supply chain management of local small and medium enterprises (SME) construction shops. We use the McKinsey 7S model for the conception of this study by providing 400 questionnaires to participants and we also used focus groups for the management guideline. From the study we found that of the seven strategies (7S) in the model (structure, strategy, systems, styles, skil...

  2. Escherichia coli surface display of single-chain antibody VRC01 against HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lin-Xu [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Mellon, Michael [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States); Bowder, Dane [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Quinn, Meghan [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States); Shea, Danielle; Wood, Charles [Nebraska Center for Virology, Lincoln, NE (United States); School of Biological Sciences, University of Nebraska—Lincoln, Lincoln, NE 68583 (United States); Xiang, Shi-Hua, E-mail: sxiang2@unl.edu [Nebraska Center for Virology, Lincoln, NE (United States); School of Veterinary Medicine and Biomedical Sciences, Lincoln, NE (United States)

    2015-01-15

    Human immunodeficiency virus type 1 (HIV-1) transmission and infection occur mainly via the mucosal surfaces. The commensal bacteria residing in these surfaces can potentially be employed as a vehicle for delivering inhibitors to prevent HIV-1 infection. In this study, we have employed a bacteria-based strategy to display a broadly neutralizing antibody VRC01, which could potentially be used to prevent HIV-1 infection. The VRC01 antibody mimics CD4-binding to gp120 and has broadly neutralization activities against HIV-1. We have designed a construct that can express the fusion peptide of the scFv-VRC01 antibody together with the autotransporter β-barrel domain of IgAP gene from Neisseria gonorrhoeae, which enabled surface display of the antibody molecule. Our results indicate that the scFv-VRC01 antibody molecule was displayed on the surface of the bacteria as demonstrated by flow cytometry and immunofluorescence microscopy. The engineered bacteria can capture HIV-1 particles via surface-binding and inhibit HIV-1 infection in cell culture. - Highlights: • Designed single-chain VRC01 antibody was demonstrated to bind HIV-1 envelope gp120. • Single-chain VRC01 antibody was successfully displayed on the surface of E. coli. • Engineered bacteria can absorb HIV-1 particles and prevent HIV-1 infection in cell culture.

  3. Escherichia coli surface display of single-chain antibody VRC01 against HIV-1 infection

    International Nuclear Information System (INIS)

    Human immunodeficiency virus type 1 (HIV-1) transmission and infection occur mainly via the mucosal surfaces. The commensal bacteria residing in these surfaces can potentially be employed as a vehicle for delivering inhibitors to prevent HIV-1 infection. In this study, we have employed a bacteria-based strategy to display a broadly neutralizing antibody VRC01, which could potentially be used to prevent HIV-1 infection. The VRC01 antibody mimics CD4-binding to gp120 and has broadly neutralization activities against HIV-1. We have designed a construct that can express the fusion peptide of the scFv-VRC01 antibody together with the autotransporter β-barrel domain of IgAP gene from Neisseria gonorrhoeae, which enabled surface display of the antibody molecule. Our results indicate that the scFv-VRC01 antibody molecule was displayed on the surface of the bacteria as demonstrated by flow cytometry and immunofluorescence microscopy. The engineered bacteria can capture HIV-1 particles via surface-binding and inhibit HIV-1 infection in cell culture. - Highlights: • Designed single-chain VRC01 antibody was demonstrated to bind HIV-1 envelope gp120. • Single-chain VRC01 antibody was successfully displayed on the surface of E. coli. • Engineered bacteria can absorb HIV-1 particles and prevent HIV-1 infection in cell culture

  4. Nanoparticles Modified With Tumor-targeting scFv Deliver siRNA and miRNA for Cancer Therapy

    Science.gov (United States)

    Chen, Yunching; Zhu, Xiaodong; Zhang, Xiaoju; Liu, Bin; Huang, Leaf

    2010-01-01

    Targeted delivery of RNA-based therapeutics for cancer therapy remains a challenge. We have developed a LPH (liposome-polycation-hyaluronic acid) nanoparticle formulation modified with tumor-targeting single-chain antibody fragment (scFv) for systemic delivery of small interfering RNA (siRNA) and microRNA (miRNA) into experimental lung metastasis of murine B16F10 melanoma. The siRNAs delivered by the scFv targeted nanoparticles efficiently downregulated the target genes (c-Myc/MDM2/VEGF) in the lung metastasis. Two daily intravenous injections of the combined siRNAs in the GC4-targeted nanoparticles significantly reduced the tumor load in the lung. miRNA-34a (miR-34a) induced apoptosis, inhibited survivin expression, and downregulated MAPK pathway in B16F10 cells. miR-34a delivered by the GC4-targeted nanoparticles significantly downregulated the survivin expression in the metastatic tumor and reduced tumor load in the lung. When miR-34a and siRNAs were co-formulated in GC4-targeted nanoparticles, an enhanced anticancer effect was observed. PMID:20606648

  5. Binding Activity Difference of Anti-CD20 scFv-Fc Fusion Protein Derived from Variable Domain Exchange

    Institute of Scientific and Technical Information of China (English)

    Shusheng Geng; Beifen Shen; Jiannan Feng; Yan Li; Yingxun Sun; Xin Gu; Ying Huang; Yugang Wang; Xianjiang Kang; Hong Chang

    2006-01-01

    Two novel engineered antibody fragments binding to antigen CD20 were generated by fusing a murine IgM-type anti-CD20 single-chain Fv fragment (scFv) to the human IgG1 CH2 (I.e., Cγ2) and CH3 (I.e., Cγ3) domains with the human IgG1 hinge (I.e. Hγ). Given the relationship between structure and function of protein, the 3-D structures of the two engineered antibody fragments were modeled using computer-aided homology modeling method.Furthermore, the relationship between 3-D conformation and their binding activity was evaluated theoretically.Due to the change of active pocket formed by CDRs, the HL23 (VH-Linker-VL-Hγ-Cγ2-Cγ3) remained its activity because of its preserved conformation, while the binding activity of the LH23 (VL-Linker-VH-Hγ-Cγ2-Cγ3) was impaired severely. Experimental studies by flow cytometry and fluorescence microscopy showed that HL23 possessed significantly superior binding activity to CD20-expressing target cells than LH23. That is to say, the order of variable regions could influence the binding activity of the fusion protein to CD20+ cell lines, which was in accordance with the theoretical results. The study highlights the potential relationship between the antibody binding activity and their 3-D conformation, which appears to be worthwhile in providing direction for future antibody design of recombinant antibody.

  6. Optimization modeling of single-chain antibody against hepatoma based on similarity algorithm.

    Science.gov (United States)

    Zhao, Zhi-Jun; Chen, Jing-Tao; Yuan, Jia-Ying; Yin, Xiao-Xiang; Song, Hua-Yong; Wang, Xin-Chun

    2015-01-01

    The purposes was to establish optimal modeling of single-chain antibody molecules based on similarity algorithm and seek the connecting peptides that had the minimal effect on the structure and bioactivity of the variable region of heavy chain (VH) and that of light chain (VL) in a single-chain antibody against liver cancer. After the Linker with different lengths (n=0~7) had been added into single chain fragment variable (ScFv), modeling of the overall sequences of VH, VL and ScFv were conducted respectively. Meanwhile, the peptide chain structure of (Gly4Ser)n was adopted for the connecting peptide. Then the spatial spherical shell layer alignment algorithm based on spherical polar coordinates was utilized for comparing the structural similarity of VH and VL before and after adding connecting peptide. Equally, in order to determine the stability of VH and VL, MATLAB was applied for analysis of the fore and aft distances and the diffusion radius. Indirect ELISA method was used to detect single-chain antibody immunological activity of Linker with different lengths. The MTT assay was utilized for the examination of the inhibition rate of single-chain antibody with different lengths of Linker to liver cancer cell. When n=4, the structural similarity between VH together with VL and their original ones was the highest. When n=3, the influence of connecting peptide on the stability of VH and VL was minimum. When n>3, the fore and aft distances changed little due to the increase and fold of the length of peptide chain. The results of ELISA detection showed that when n=4, affinity of single chain antibody to liver cancer cells was much higher. The MTT test also indicated that when n=4, the inhibition rate of the connecting peptide on hepatoma carcinoma cell reached the highest, and that came second when n=3. When n=4, the structural stability and biological functions of anti-hepatoma single-chain antibody were both favorable. This study has provided a basis for the design

  7. Clinical Applications of Phage-Derived sFvs and sFv Fusion Proteins

    Directory of Open Access Journals (Sweden)

    K. A. Chester

    2000-01-01

    Full Text Available Single chain Fv antibodies (sFvs have been produced from filamentous bacteriophage libraries obtained from immunised mice. MFE-23, the most characterised of these sFvs, is reactive with carcinoembryonic antigen (CEA, a glycoprotein that is highly expressed in colorectal adenocarcinomas. MFE-23 has been expressed in bacteria and purified in our laboratory for two clinical trials; a gamma camera imaging trial using 123I-MFE-23 and a radioimmunoguided surgery trial using 125I-MFE-23, where tumour deposits are detected by a hand-held probe during surgery. Both these trials show MFE-23 is safe and effective in localising tumour deposits in patients with cancer. We are now developing fusion proteins which use MFE-23 to deliver a therapeutic moiety; MFE-23::CPG2 targets the enzyme carboxypeptidase G2 (CPG2 for use in the ADEPT (antibody directed enzyme prodrug therapy system and MFE::TNFα aims to reduce sequestration and increase tumor concentrations of systemically administered TNFα.

  8. Characterization of a single-chain variable fragment recognizing a linear epitope of aβ: a biotechnical tool for studies on Alzheimer's disease?

    Directory of Open Access Journals (Sweden)

    Silke Dornieden

    Full Text Available Alzheimer's disease (AD is a progressive neurodegenerative disorder with devastating effects. Currently, therapeutic options are limited to symptomatic treatment. For more than a decade, research focused on immunotherapy for the causal treatment of AD. However, clinical trials with active immunization using Aβ encountered severe complications, for example meningoencephalitis. Consequently, attention focused on passive immunization using antibodies. As an alternative to large immunoglobulins (IgGs, Aβ binding single-chain variable fragments (scFvs were used for diagnostic and therapeutic research approaches. scFvs can be expressed in E. coli and may provide improved pharmacokinetic properties like increased blood-brain barrier permeability or reduced side-effects in vivo. In this study, we constructed an scFv from an Aβ binding IgG, designated IC16, which binds the N-terminal region of Aβ (Aβ(1-8. scFv-IC16 was expressed in E. coli, purified and characterized with respect to its interaction with different Aβ species and its influence on Aβ fibril formation. We were able to show that scFv-IC16 strongly influenced the aggregation behavior of Aβ and could be applied as an Aβ detection probe for plaque staining in the brains of transgenic AD model mice. The results indicate potential for therapy and diagnosis of AD.

  9. Click ‘Like’ and Post It on Your Wall! Chain Posts on Facebook – Identity Construction and Values

    OpenAIRE

    Piret Voolaid

    2013-01-01

    The article focuses on chain posts that were collected in the years 2010–2012 and spread predominantly among girls of ten to twelve on Facebook (facebook.com) – a social network that has a membership of over 450,000 in Estonia. The source material comprising approximately 220 texts is similar by form and content to chain letters known from earlier tradition; yet, the web environment with its specific technical structure allows the texts to turn into a peculiar Facebook-like phenomenon.The aut...

  10. Effects of tacrolimus on infection of Friend murine leukemia virus to Fv-4 gene heterozygous mice

    Institute of Scientific and Technical Information of China (English)

    Feng-min ZHANG; Bao-feng YANG; Hong-xi GU; Xiao-bei CHEN; Zhao-hua ZHONG; Zhi CHENG

    2004-01-01

    AIM: To investigate the effect of tacrolimus (FK506) on the infection of Friend murine leukemia virus (Friend MuLV) in vivo. METHODS: Three kinds of mice were used including Friend MuLV-sensitive BALB/c mice, Friend MuLV-resistant Fv-4 gene-homozygous mice (Fv-4 mice), and Friend MuLV-resistant Fv-4 gene-heterozygous mice (Fl mice). Tacrolimus was administrated ip to those mice in every 2 d. Those treated mice were inoculated ip with Friend MuLV once on d 3. The symptoms and viral proliferations in those mice were observed to recognize the Friend MuLV infection. The expression and genotype of Fv-4 gene that resistant against the infection of Friend MuLV were analyzed to confirm the genomic background and related mechanism of the resistance. RESULTS:BALB/c mice and Fl mice, but not Fv-4 mice, appeared obvious early death, spleenomegaly, and viral proliferation after both treatments of viral inoculation and tacrolimus administration, whereas the expression and genotype of Fv-4 gene was not changed in F1 mice and Fv-4 mice with treatment of tacrolimus. Compared to the virusinoculated control, the Friend MuLV-sensitivity of tacrolimus-treated BALB/c mice and the Friend MuLV-resistance of tacrolimus-treated Fv-4 mice were the same as the controls, but only Fl mice became the symptoms and viral proliferation after both treatments. It suggested the Friend MuLV-resistant Fl mice could be converted to be Friend MuLV-sensitive by treatment of tacrolimus, and this conversion was not depended on the expression and genotype of Fv-4 gene. CONCLUSION: Tacrolimus could not inhibit the infection of Friend MuLV in all mice, furthermore,it could enhance the infection of Friend MuLV in F1 mice. The enhancement may be related to the immunosuppressive effect of tacrolimus.

  11. Preparation and Identification of a Single-chain Variable Fragment Antibody Against Canine Distemper Virus.

    Science.gov (United States)

    Yi, Li; Cheng, Shipeng

    2015-08-01

    The variable regions of the heavy chain (VH) and light chain (VL) were amplified by RT-PCR from the hybridoma 1N8, which secretes the monoclonal antibody against CDV N protein (aa 277-471). The VL and VH amplicons were combined using SOE-PCR by a 12 amino acid flexible linker (SSGGGGSGGGGS), which produced the scFv gene (named scFv/1N8). After sequence analysis, the scFv/1N8 gene was cloned into the prokaryotic expression vector PET32a with a His-tag. The recombinant scFv/1N8 protein was successfully expressed in recombinant Escherichia coli by IPTG induction. Moreover, the binding activity and specificity of the scFv were determined by indirect ELISA (His-tag) and competitive ELISA. The recombinant scFv/1N8 protein reported here will provide some basis for further antiviral drug research based on the scFv molecule. PMID:26301925

  12. Aptamers, antibody scFv, and antibody Fab' fragments: An overview and comparison of three of the most versatile biosensor biorecognition elements.

    Science.gov (United States)

    Crivianu-Gaita, Victor; Thompson, Michael

    2016-11-15

    The choice of biosensing elements is crucial for the development of the optimal biosensor. Three of the most versatile biosensing elements are antibody single-chain Fv fragments (scFv), antibody fragment-antigen binding (Fab') units, and aptamers. This article provides an overview of these three biorecognition elements with respects to their synthesis/engineering, various immobilization techniques, and examples of their use in biosensors. Furthermore, the final section of the review compares and contrasts their characteristics (time/cost of development, ease and variability of immobilization, affinity, stability) illustrating their advantages and disadvantages. Overall, scFv fragments are found to display the highest customizability (i.e. addition of functional groups, immobilizing peptides, etc.) due to recombinant synthesis techniques. If time and cost are an issue in the development of the biosensor, Fab' fragments should be chosen as they are relatively cheap and can be developed quickly from whole antibodies (several days). However, if there are sufficient funds and time is not a factor, aptamers should be utilized as they display the greatest affinity towards their target analytes and are extremely stable (excellent biosensor regenerability). PMID:27155114

  13. Antitumor activities of dFv-LDP-AE: An enediyne-energized fusion protein targeting tumor-associated antigen gelatinases.

    Science.gov (United States)

    Zhong, Gen-Shen; Wu, Min-Na; Guo, Xiao-Fang; Zhang, Sheng-Hua; Miao, Qing-Fang; Zhen, Yong-Su

    2012-10-01

    Gelatinases play an important role in tumor growth and metastasis, and overexpression of these molecules is strongly correlated with poor prognosis in a variety of malignant tumors. Lidamycin is an enediyne antitumor antibiotic with potent cytotoxicity. We previously reported that a tandem scFv format (dFv-LDP-AE) showed enhanced binding ability with gelatinases compared with the scFv-lidamycin conjugate (Fv-LDP-AE). In this study, the antitumor activities of dFv-LDP-AE on hepatocellular carcinoma (HCC) were evaluated in vitro and in vivo. By SDS-PAGE analysis, it was found that partial fusion protein dFv-LDP existed as dimer; the results of ELISA and immunofluorescence demonstrated that the fusion protein dFv-LDP could efficiently bind to hepatoma cells in vitro. The apparent arrest of cell cycle at G2/M phase and induction of apoptosis at nanomole levels indicated that the dFv-LDP-AE was very potent against HCC. In in vivo experiments, dFv-LDP-AE shown enhanced cytotoxic effects compared to those of LDM. Administration at mouse tolerable dosage level, the inhibition rate of tumor growth was 89.5% of dFv-LDP-AE vs. 73.6% of LDM on transplantable H22 in mice (Penediyne-energized fusion protein dFv-LDP-AE showed potential application as a new agent for therapeutic appications in HCC. PMID:22797730

  14. 关于医药零售连锁企业信息化建设的探讨%Probe into the Informatization Construction of Pharmaceutical Retail Chains

    Institute of Scientific and Technical Information of China (English)

    剧晓红

    2015-01-01

    This paper expounds the current status of the informatization construction of pharmaceutical retail chains, analyzes the benefits brought about by the informatization construction to enterprises, puts forward some improvement countermeasures, and based on this, briefly probes into the development trends of the informatization construction of pharmaceutical retail chains in the aspects of cloud computing, pharmaceutical e-commerce, and new media, etc. in the Internet era.%阐述了医药零售连锁企业信息化建设的现状,分析了信息化建设给企业带来的效益,提出了改进对策,并在此基础上简单探讨了互联网时代医药零售连锁企业信息化建设在云计算、医药电子商务、新媒体等方面的发展趋势。

  15. Supply chain for new construction: buy where we build; Cadena de suministro para las nuevas construcciones: Buy where we build

    Energy Technology Data Exchange (ETDEWEB)

    Cruz, J. L.; Bueno, S.

    2012-07-01

    A global supply strategy has been placed by Westinghouse in order to face the current and future constructions. And in this way, Westinghouse will be able to take advantage of benefit of Local Supply. this article shows the Westinghouse Global Supply Strategy, Local Supply considerations, the current state of supplier calcifications, and finally Memorandums of Understanding and Alliances agreed with local suppliers around the world to provide a suitable solution for the new construction needs. (Author)

  16. Study on Brand Construction from the Perspective of Supply Chain Management%基于供应链视角的品牌建设研究

    Institute of Scientific and Technical Information of China (English)

    宋爱华

    2016-01-01

    Brand is an important embodiment of the enterprise and even the country's competitive power. Now It has entered the era of brand competition and brand consumption,the brand is more and more become the focus of popular attention. Who can walk in the forefront of the brand construction,create a famous brand and a strong brand,who will be able to win the competition. But the traditional brand-building ideas ignored the supply chain management issues,enterprises should think about the strategy of brand construction from the perspective of supply chain management,strengthen the effective control of supply chain operation,act brand construction as a strategic means to improve the quality of the supply chain operation,enhance the competitive ability of the supply chain,thus can cultivate a famous brand and a strong brand,make sure the enterprise last for a long time.%品牌是一个企业乃至国家竞争实力的重要体现。现在已经进入到品牌竞争和品牌消费的时代,品牌越来越成为人们关注的焦点。谁能够走在品牌建设的前列,打造出知名品牌、强势品牌,谁就能够赢得竞争,但传统的品牌建设思路忽视了供应链管理问题,应从供应链管理的角度思考品牌建设的策略,加强供应链运行有效管控,以品牌建设为战略手段提升供应链运行的质量,增强供应链的竞争能力,才能够培育打造出知名品牌、强势品牌,保证企业持续长久。

  17. Need of a consistent and convenient nucleus identification in ENDF files for the automatic construction of the depletion chains

    Directory of Open Access Journals (Sweden)

    Mosca Pietro

    2016-01-01

    As a main result of this work, our suggestion is to change the ENDF format using systematically the isomeric state number to identify the nuclei. This proposal is already compliant to a huge amount ENDF data that are not in agreement with the present ENDF format. This choice is the most convenient because, ultimately, it allows one to give human readable names to the nuclei of the depletion chains.

  18. How to measure Innovative Supply Chain Strategies (ISCS) in the context of SCM? -Construct development and measurement validation

    OpenAIRE

    Lavastre, Olivier; AGERON, Blandine; Chaze-Magnan, L.

    2014-01-01

    Nowadays, organizations have to innovate either in product or in management (Damanpour and Aravind, 2012). Ifinnovations in product are largely studied (Garcia and Calantone, 2002), managerial innovations are seldom studied(Soosay et al., 2008; Arlbjørn et al., 2011). Simultaneously, organizations develop Supply Chain Management(SCM) to manage their inter-organizational relationships with their partners (Chen and Paulraj, 2004). SCM“constitutes an interesting potential area for creating compe...

  19. The antitumor efficacy of a novel adenovirus-mediated anti-p21Ras single chain fragment variable antibody on human cancers in vitro and in vivo.

    Science.gov (United States)

    Yang, Ju-Lun; Pan, Xin-Yan; Zhao, Wen-Xing; Hu, Qi-Chan; Ding, Feng; Feng, Qiang; Li, Gui-Yun; Luo, Ying

    2016-03-01

    Activated ras genes are found in a large number of human tumors, and therefore are one of important targets for cancer therapy. This study investigated the antitumor effects of a novel single chain fragment variable antibody (scFv) against ras protein, p21Ras. The anti-p21Ras scFv gene was constructed by phage display library from hybridoma KGHR1, and then subcloned into replication-defective adenovirus vector to obtain recombinant adenovirus KGHV100. Human tumor cell lines with high expression of p21Ras SW480, MDA-MB‑231, OVCAR-3, BEL-7402, as well as tumor cell line with low expression of p21Ras, SKOV3, were employed to investigate antitumor effects in vitro and in vivo. Fluorescence microscopy demonstrated that KGHV100 was able to express intracellularly anti-p21Ras scFv antibody in cultured tumor cells and in transplantation tumor cells. MTT, Transwell, colony formation, and flow cytometry analysis showed that KGHV100 led to significant growth arrest in tumor cells with high p21Ras expression, and induced G0/G1 cell cycle arrest in the studied tumor cell lines. In vivo, KGHV100 significantly inhibited tumor growth following intratumoral injection, and the survival rates of the mice were higher than the control group. These results indicate that the adenovirus-mediated intracellular expression of the novel anti-p21Ras scFv exerted strong antitumoral effects, and may be a potential method for therapy of cancers with p21Ras overexpression. PMID:26780944

  20. AFSC/ABL: Origins of salmon seized from the F/V Arctic Wind

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Samples of chum (Oncorhynchus keta), sockeye (O. nerka), and chinook salmon (O. tshawytscha) seized from the F/V Arctic Wind were analyzed to determine their region...

  1. Expression of recombinant human anti-TNF-α scFv-Fc in Arabidopsis thaliana seeds.

    Science.gov (United States)

    Yao, N; Ai, L; Dong, Y Y; Liu, X M; Wang, D Z; Wang, N; Li, X W; Wang, F W; Li, Xk; Li, H Y; Jiang, C

    2016-01-01

    Recombinant human anti-tumor necrosis factor (TNF)-α scFv-Fc was expressed in TKO mutant Arabidopsis thaliana seeds using plant-specific codons. Immunoblotting using a human IgG1 antibody detected the expression of anti-TNF-α proteins in plants. Results from qRT-PCR analysis demonstrated that the time of harvest significantly affected the protein yield and quality. Our results indicate that the Phaseolus vulgaris β-phaseolin promoter directed anti-TNF-α scFv-Fc expression in A. thaliana seeds, with a maximum yield obtained at 20-days of development. Although the yield of anti-TNF-α scFv-Fc protein was not very high, accumulation of recombinant proteins in seeds is an attractive and simple method that can be used to purify biologically active anti-TNF-α scFv-Fc. PMID:27420937

  2. McKinsey 7S Model for Supply Chain Management of Local SMEs Construction Business in Upper Northeast Region of Thailand

    Directory of Open Access Journals (Sweden)

    Thanaphan Naipinit

    2014-03-01

    Full Text Available The purpose of this study is to study the successful business strategies and the guidelines for the management strategies of supply chain management of local small and medium enterprises (SME construction shops. We use the McKinsey 7S model for the conception of this study by providing 400 questionnaires to participants and we also used focus groups for the management guideline. From the study we found that of the seven strategies (7S in the model (structure, strategy, systems, styles, skill, staff, and shared values, most entrepreneurs scored highly in strategy; however, most entrepreneurs scored low in several areas: working with software applications, lack of outside training, and most entrepreneurs maintain command as the owner and do not give authority to others. In addition, the Thai government should create a policy by collaborating with Lao PDR to reduce some barriers of international trade to help SME construction shops in Thailand.

  3. Production and characterization of a single-chain variable fragment linked alkaline phosphatase fusion protein for detection of O,O-diethyl organophosphorus pesticides in a one-step enzyme-linked immunosorbent assay

    Science.gov (United States)

    A single-chain variable fragment (scFv) and alkaline phosphatase (AP) fusion protein for detection of O, O-diethyl organophosphorus pesticides (OPs) was produced and characterized. The scFv gene was prepared by cloning VL and VH genes from a hybridoma cell secreting monoclonal antibody with broad-s...

  4. Construction, Expression and Characterization of a Chimeric Protein Targeting Carcinoembryonic Antigen in Lung Cancer

    Institute of Scientific and Technical Information of China (English)

    LI Yang; HUA Shu-cheng; MA Cheng-yuan; YU Zhen-xiang; XU Li-jun; LI Dan; SUN Li-li; LI Xiao; PENG Li-ping

    2011-01-01

    The carcinoembryonic antigen(CEA) is an oncofetal glycoprotein known as an important clinical tumor marker and is overexpressed in several types of tumors, including colorectal and lung carcinomas. We constructed a chimeric protein that exhibits both specific binding and immune stimulating activities, by fusing staphylococcal enterotoxin A(SEA) to the C-terminus of an anti-CEA single-chain disulfide-stabilized Fv(scdsFv) antibody (single-chain-C-terminus/SEA, SC-C/SEA). The SC-C/SEA protein was expressed in Escherichia coli(E. coli), refolded, and purified on an immobilized Ni2+ affinity chromatography column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) and Western blot analysis reveal that the target protein was expressed sufficiently. We used immunofluorescence assays to demonstrate that SC-C/SEA could bind specifically to human lung carcinoma cells(A549), but almost human uterine cervix cells(HeLa). We also used the L-lactate dehydrogenase(LDH) release assay to show that SC-C/SEA elicits a strong A549 tumor-specific cytotoxic T lymphocyte(CTL) response in vitro. The results suggest that SC-C/SEA shows specific activity against CEA-positive cells and has potential application in CEA-targeted cancer immunotherapy.

  5. Preparation of a Chicken scFv to Analyze Gentamicin Residue in Animal Derived Food Products.

    Science.gov (United States)

    Li, Cui; He, Jinxin; Ren, Hao; Zhang, Xiaoying; Du, Enqi; Li, Xinping

    2016-04-01

    Chicken is an ideal model for simplified recombinant antibody library generation. It has been rarely been reported to apply chicken single-chain variable fragments (scFvs) in immunoassays for the detection of antibiotic and chemical contaminants in animal food products. In this study, the scFvs (S-1 and S-5) were isolated from a phage display library derived from a hyperimmunized chicken. The checker board titration revealed that the optimum concentrations of S-1 and S-5 were 0.78 μg/mL and 0.44 μg/mL respectively, to obtain OD450 around 1.0 at 5 μg/mL of Gent-OVA coating concentration. Both S-1 and S-5 exhibited negligible cross reactivity with kanamycin and amikacin. The 50% inhibitory concentration (IC50) of S-1 and S-5 were 12.418 ng/mL and 14.674 ng/mL respectively. In the indirect competitive ELISA (ic-ELISA), the limits of detection for S-1 and S-5 were 0.147 ng/mL and 0.219 ng/mL respectively. The mean recovery for Gent ranged from 60.91% to 118.09% with no more than 10.35% relative standard deviation (RSD) between the intra-assay and the inter-assay. These results indicate the chicken scFv based ic-ELISA method is suitable for the detection of Gent residue in animal derived edible tissues and milk. PMID:26980703

  6. Human scFv antibodies (Afribumabs) against Africanized bee venom: Advances in melittin recognition.

    Science.gov (United States)

    Pessenda, Gabriela; Silva, Luciano C; Campos, Lucas B; Pacello, Elenice M; Pucca, Manuela B; Martinez, Edson Z; Barbosa, José E

    2016-03-15

    Africanized Apis mellifera bees, also known as killer bees, have an exceptional defensive instinct, characterized by mass attacks that may cause envenomation or death. From the years 2000-2013, 77,066 bee accidents occurred in Brazil. Bee venom comprises several substances, including melittin and phospholipase A2 (PLA2). Due to the lack of antivenom for bee envenomation, this study aimed to produce human monoclonal antibody fragments (single chain fragment variable; scFv), by using phage display technology. These fragments targeted melittin and PLA2, the two major components of bee venom, to minimize their toxic effects in cases of mass envenomation. Two phage antibody selections were performed using purified melittin. As the commercial melittin is contaminated with PLA2, phages specific to PLA2 were also obtained during one of the selections. Specific clones for melittin and PLA2 were selected for the production of soluble scFvs, named here Afribumabs: prefix: afrib- (from Africanized bee); stem/suffix: -umab (fully human antibody). Afribumabs 1 and 2 were tested in in vitro and in vivo assays to assess their ability to inhibit the toxic actions of purified melittin, PLA2, and crude bee venom. Afribumabs reduced hemolysis caused by purified melittin and PLA2 and by crude venom in vitro and reduced edema formation in the paws of mice and prolonged the survival of venom-injected animals in vivo. These results demonstrate that Afribumabs may contribute to the production of the first non-heterologous antivenom treatment against bee envenomation. Such a treatment may overcome some of the difficulties associated with conventional immunotherapy techniques. PMID:26829652

  7. Anti-HER2 antibody and ScFvEGFR-conjugated antifouling magnetic iron oxide nanoparticles for targeting and magnetic resonance imaging of breast cancer

    Directory of Open Access Journals (Sweden)

    Chen H

    2013-10-01

    Full Text Available Hongwei Chen,1,* Liya Wang,1,2,* Qiqi Yu,1,2 Weiping Qian,3 Diana Tiwari,1 Hong Yi,4 Andrew Y Wang,5 Jing Huang,1,2 Lily Yang,3 Hui Mao1,2 1Department of Radiology and Imaging Sciences, 2Center for Systems Imaging, 3Department of Surgery, Emory University School of Medicine, 4Robert Apkarian Electron Microscopy Core, Emory University, Atlanta, GA, 5Ocean NanoTech LLC, Springdale, AK, USA *These authors contributed equally to this work Abstract: Antifouling magnetic iron oxide nanoparticles (IONPs coated with block copolymer poly(ethylene oxide-block-poly(γ-methacryloxypropyltrimethoxysilane (PEO-b-PγMPS were investigated for improving cell targeting by reducing nonspecific uptake. Conjugation of a HER2 antibody, Herceptin®, or a single chain fragment (ScFv of antibody against epidermal growth factor receptor (ScFvEGFR to PEO-b-PγMPS-coated IONPs resulted in HER2-targeted or EGFR-targeted IONPs (anti-HER2-IONPs or ScFvEGFR-IONPs. The anti-HER2-IONPs bound specifically to SK-BR-3, a HER2-overexpressing breast cancer cell line, but not to MDA-MB-231, a HER2-underexpressing cell line. On the other hand, the ScFvEGFR-IONPs showed strong reactivity with MDA-MB-231, an EGFR-positive human breast cancer cell line, but not with MDA-MB-453, an EGFR-negative human breast cancer cell line. Transmission electron microscopy revealed internalization of the receptor-targeted nanoparticles by the targeted cancer cells. In addition, both antibody-conjugated and non-antibody-conjugated IONPs showed reduced nonspecific uptake by RAW264.7 mouse macrophages in vitro. The developed IONPs showed a long blood circulation time (serum half-life 11.6 hours in mice and low accumulation in both the liver and spleen. At 24 hours after systemic administration of ScFvEGFR-IONPs into mice bearing EGFR-positive breast cancer 4T1 mouse mammary tumors, magnetic resonance imaging revealed signal reduction in the tumor as a result of the accumulation of the targeted IONPs

  8. Suppression of Aggrus/podoplanin-induced platelet aggregation and pulmonary metastasis by a single-chain antibody variable region fragment

    International Nuclear Information System (INIS)

    Almost all highly metastatic tumor cells possess high platelet aggregating abilities, thereby form large tumor cell-platelet aggregates in the microvasculature. Embolization of tumor cells in the microvasculature is considered to be the first step in metastasis to distant organs. We previously identified the platelet aggregation-inducing factor expressed on the surfaces of highly metastatic tumor cells and named as Aggrus. Aggrus was observed to be identical to the marker protein podoplanin (alternative names, T1α, OTS-8, and others). Aggrus is frequently overexpressed in several types of tumors and enhances platelet aggregation by interacting with the platelet receptor C-type lectin-like receptor 2 (CLEC-2). Here, we generated a novel single-chain antibody variable region fragment (scFv) by linking the variable regions of heavy and light chains of the neutralizing anti-human Aggrus monoclonal antibody MS-1 with a flexible peptide linker. Unfortunately, the generated KM10 scFv failed to suppress Aggrus-induced platelet aggregation in vitro. Therefore, we performed phage display screening and finally obtained a high-affinity scFv, K-11. K-11 scFv was able to suppress Aggrus-induced platelet aggregation in vitro. Moreover, K-11 scFv prevented the formation of pulmonary metastasis in vivo. These results suggest that K-11 scFv may be useful as metastasis inhibitory scFv and is expected to aid in the development of preclinical and clinical examinations of Aggrus-targeted cancer therapies

  9. Characterization of antibodies in single-chain format against the E7 oncoprotein of the Human papillomavirus type 16 and their improvement by mutagenesis

    International Nuclear Information System (INIS)

    Human papillomaviruses (HPV) are the etiological agents of cervical cancer. The viral E7 protein plays a crucial role in viral oncogenesis. Many strategies have been explored to block the E7 oncoprotein activity. The single-chain variable antibody fragments (scFvs) are valuable tools in cancer immunotherapy and can be used as 'intracellular antibodies' to knock out specific protein functions. For both in vivo and in vitro employment, the scFv intrinsic solubility and stability are important to achieve long-lasting effects. Here we report the characterization in terms of reactivity, solubility and thermal stability of three anti-HPV16 E7 scFvs. We have also analysed the scFv43 sequence with the aim of improving stability and then activity of the antibody, previously shown to have antiproliferative activity when expressed in HPV16-positive cells. The three anti-HPV16 E7 scFv 32, 43 51 were selected from the ETH-2 'phage-display' library. Thermal stability was evaluated with ELISA by determining the residual activity of each purified scFv against the recombinant HPV16 E7, after incubation in the presence of human seroalbumine for different time-intervals at different temperatures. Sequence analysis of the scFvs was performed with BLAST and CLUSTALL programs. The scFv43 aminoacid changes were reverted back to the consensus sequence from the immunoglobuline database by site-directed mutagenesis. ScFv solubility was evaluated with Western blotting by determining their relative amounts in the soluble and insoluble fractions of both prokaryotic and eukaryotic systems. ScFv51 was the most thermally stable scFv considered. Sequence analysis of the most reactive scFv43 has evidenced 2 amino acid changes possibly involved in molecule stability, in the VH and VL CDR3 regions respectively. By mutagenesis, two novel scFv43-derived scFvs were obtained, scFv43 M1 and M2. ScFv43 M2 showed to have improved thermal stability and solubility in comparison with the

  10. Characterization of antibodies in single-chain format against the E7 oncoprotein of the Human papillomavirus type 16 and their improvement by mutagenesis

    Directory of Open Access Journals (Sweden)

    Accardi Luisa

    2007-01-01

    Full Text Available Abstract Background Human papillomaviruses (HPV are the etiological agents of cervical cancer. The viral E7 protein plays a crucial role in viral oncogenesis. Many strategies have been explored to block the E7 oncoprotein activity. The single-chain variable antibody fragments (scFvs are valuable tools in cancer immunotherapy and can be used as "intracellular antibodies" to knock out specific protein functions. For both in vivo and in vitro employment, the scFv intrinsic solubility and stability are important to achieve long-lasting effects. Here we report the characterization in terms of reactivity, solubility and thermal stability of three anti-HPV16 E7 scFvs. We have also analysed the scFv43 sequence with the aim of improving stability and then activity of the antibody, previously shown to have antiproliferative activity when expressed in HPV16-positive cells. Methods The three anti-HPV16 E7 scFv 32, 43 51 were selected from the ETH-2 "phage-display" library. Thermal stability was evaluated with ELISA by determining the residual activity of each purified scFv against the recombinant HPV16 E7, after incubation in the presence of human seroalbumine for different time-intervals at different temperatures. Sequence analysis of the scFvs was performed with BLAST and CLUSTALL programs. The scFv43 aminoacid changes were reverted back to the consensus sequence from the immunoglobuline database by site-directed mutagenesis. ScFv solubility was evaluated with Western blotting by determining their relative amounts in the soluble and insoluble fractions of both prokaryotic and eukaryotic systems. Results ScFv51 was the most thermally stable scFv considered. Sequence analysis of the most reactive scFv43 has evidenced 2 amino acid changes possibly involved in molecule stability, in the VH and VL CDR3 regions respectively. By mutagenesis, two novel scFv43-derived scFvs were obtained, scFv43 M1 and M2. ScFv43 M2 showed to have improved thermal stability and

  11. Recombinant anti-tenascin antibody constructs

    Energy Technology Data Exchange (ETDEWEB)

    ZALUTSKY, MICHAEL R

    2006-08-29

    -tenascin constructs with optimized properties for use in tandem with short half life radionuclides such as 211At ( as well as 1.8-hr 18F for PET imaging) is warranted. Our specific aims are: 1) to construct a bivalent, anti-tenascin molecule containing murine 81C6 variable regions and the human IgG2 hinge region. Both the CH2 domain deletion construct (CH2) and F(ab’)2 will be investigated; 2) to construct a single-chain Fv dimer or multimer with adequate stability, affinity and immunoreactivity for use in tandem with 211At for therapy and 18F for imaging; 3) to generate higher affinity scFv constructs reactive with the alternatively spliced fibronectin type III repeats CD of the tenascin molecule via phage display technology and site-directed mutagenesis; 4) to label promising anti-tenascin constructs with radioiodine, 211At, and 18F and evaluate their potential as radiodiagnostic and radiotherapeutic agents. The proposed studies include: characterization of affinity and immunoreactivity after labeling; evaluation of tissue distribution and projected dosimetry in normal mice, and athymic rodents with subcutaneous, intracranial and neoplastic meningitis xenografts; investigation of the nature of low and high molecular weight labeled catabolites generated in mice; and assessment of cytotoxicity in vitro and in vivo models of human glioma, and possibly, other tenascin expressing tumors; and 5) to investigate strategies for labeling scFv monomers and dimers which will minimize retention of the radiohalogen in the kidneys through the use of negatively charged templates.

  12. Annual parallax and a dimming event of a Mira variable star, FV Bootis

    Science.gov (United States)

    Kamezaki, Tatsuya; Nakagawa, Akiharu; Omodaka, Toshihiro; Inoue, Kan-ichiro; Chibueze, James O.; Nagayama, Takumi; Ueno, Yuji; Matsunaga, Noriyuki

    2016-08-01

    We present the first measurement of the trigonometric parallax of water masers associated with a Mira star, FV Bootis (FV Boo) using VLBI Exploration of Radio Astrometry (VERA). Based on our multi-epoch VERA observations, we derived the parallax to be 0.97 ± 0.06 mas, which corresponds to a distance of 1.03^{+0.07}_{-0.06} kpc. The water masers around FV Boo were spatially distributed over an area of 41 au × 41 au, and their internal motions indicate the presence of an outflow. Using the Kagoshima University 1 m optical/infrared telescope, we determined the period to be 305.6 d and the mean apparent magnitude to be +2.91 mag in the K'-band. On the period-luminosity plane, the obtained period and K'-band magnitude puts FV Boo slightly below the sequence of Miras, possibly due to circumstellar reddening. Combining our photometric data with COBE and 2MASS datasets spanning over 20 years, we found in the near infrared that FV Boo was significantly fainter in 2005 compared with preceding and later phases. Its color, however, did not show a large variation through this change. We infer that the dimming could be caused by an eclipse due to a cloud in a binary system.

  13. Modular Synthetic Platform for the Construction of Functional Single-Chain Polymeric Nanoparticles: From Aqueous Catalysis to Photosensitization.

    Science.gov (United States)

    Liu, Yiliu; Pauloehrl, Thomas; Presolski, Stanislav I; Albertazzi, Lorenzo; Palmans, Anja R A; Meijer, E W

    2015-10-14

    Single-chain polymeric nanoparticles (SCPNs) are intriguing systems for multiple applications. In order to arrive at a controlled, but random, positioning of the different side groups to the polymer backbone, alternative synthetic routes have to be developed. Here, a general postpolymerization modification strategy of poly(pentafluorophenyl acrylate) (pPFPA) is presented as a versatile method to rapidly access functional SCPNs. We first show that the sequential addition of a benzene-1,3,5-tricarboxamide-based amine, acting as the supramolecular recognition motif, and water-soluble polyetheramine (Jeffamine) to pPFPA affords random copolymers that fold in water into SCPNs. The scope of the modular platform is illustrated by preparing two types of functional SCPNs. First, we prepared SCPNs designed for bio-orthogonal catalysis by attaching pendant mono(benzimidazoylmethyl)-bis(pyridylmethyl) (Bimpy), phenanthroline (Phen), or 2,2'-bipyridine (BiPy), ligands capable of binding either Cu(I) or Pd(II). The Bimpy- and Phen-containing SCPNs ligated to Cu(I) significantly accelerate azide-alkyne cycloaddition reactions while Bipy-containing SCPNs ligated to Pd(II) efficiently catalyze depropargylation reactions. In all cases, reactions proceeded efficiently in phosphate buffer at a physiological pH and at low substrate concentrations. Next, the potential of SCPNs for photodynamic therapy was evaluated. Introducing porphyrins in SCPNs leads to novel photosensitizers that can produce singlet oxygen ((1)O2) upon photoirradiation. Additionally, by attaching both porphyrins and prodrug models, attached via (1)O2-cleavable amino-acrylate linker, to the SCPNs, we show that irradiation of the SCPNs results in a cascade reaction of (1)O2 generation followed by cleavage of the amino-acrylate linkers, releasing the drug model. The modular synthesis strategy reported here provides rapid and controlled access to SCPNs with tunable amounts of active units that fulfill different

  14. Fields From Markov Chains

    DEFF Research Database (Denmark)

    Justesen, Jørn

    2005-01-01

    A simple construction of two-dimensional (2-D) fields is presented. Rows and columns are outcomes of the same Markov chain. The entropy can be calculated explicitly.......A simple construction of two-dimensional (2-D) fields is presented. Rows and columns are outcomes of the same Markov chain. The entropy can be calculated explicitly....

  15. Detection of activated platelets in a mouse model of carotid artery thrombosis with 18F-labeled single-chain antibodies

    International Nuclear Information System (INIS)

    Introduction: Activated platelets are key players in thrombosis and inflammation. We previously generated single-chain antibodies (scFv) against ligand-induced binding sites (LIBS) on the highly abundant platelet glycoprotein integrin receptor IIb/IIIa. The aim of this study was the construction and characterisation of a novel 18F PET radiotracer based on this antibody. Methods: ScFvanti-LIBS and control antibody mut-scFv were reacted with N-succinimidyl-4-[18F]fluorobenzoate (S[18F]FB). Radiolabeled scFv was incubated with in vitro formed platelet clots and injected into mice with FeCl3 induced thrombus in the left carotid artery. Clots were imaged in the PET scanner and amount of radioactivity measured using an ionization chamber and image analysis. Assessment of vessel injury as well as the biodistribution of the radiolabeled scFv was studied. Results: After incubation with increasing concentrations of 18F-scFvanti-LIBS clots had retained significantly higher amounts of radioactivity compared to clots incubated with radiolabeled 18F-mut-scFv (13.3 ± 3.8 vs. 3.6 ± 1 KBq, p < 0.05, n = 9, decay corrected). In the in vivo experiments we found an high uptake of the tracer in the injured vessel compared with the non-injured vessel, with 12.6 ± 4.7% injected dose per gram (ID/g) uptake in the injured vessel and 3.7 ± 0.9% ID/g in the non-injured vessel 5 minutes after injection (p < 0.05, n = 6). Conclusions: Our results show that the novel antibody radiotracer 18F-scFvanti-LIBS is useful for the sensitive detection of activated platelets and thrombosis. Advances in knowledge and implications for patient care: We describe the first 18F variant of a scFvanti-LIBS against activated platelets. This diagnostic agent could provide a powerful tool for the assessment of acute thrombosis and inflammation in patients in the future

  16. Dynamics of Apis mellifera Filamentous Virus (AmFV Infections in Honey Bees and Relationships with Other Parasites

    Directory of Open Access Journals (Sweden)

    Ulrike Hartmann

    2015-05-01

    Full Text Available Apis mellifera filamentous virus (AmFV is a large double stranded DNA virus of honey bees, but its relationship with other parasites and prevalence are poorly known. We analyzed individual honey bees from three colonies at different times post emergence in order to monitor the dynamics of the AmFV gut colonization under natural conditions. Prevalence and loads of microsporidia and trypanosomes were also recorded, as well as five common honey bee RNA viruses. The results show that a high proportion of bees get infected with AmFV during the first week post-emergence (75% and that AmFV DNA levels remained constant. A similar pattern was observed for microsporidia while trypanosomes seem to require more time to colonize the gut. No significant associations between these three infections were found, but significant positive correlations were observed between AmFV and RNA viruses. In parallel, the prevalence of AmFV in France and Sweden was assessed from pooled honey bee workers. The data indicate that AmFV is almost ubiquitous, and does not seem to follow seasonal patterns, although higher viral loads were significantly detected in spring. A high prevalence of AmFV was also found in winter bees, without obvious impact on overwintering of the colonies.

  17. Radioiodination and biodistribution of the monoclonal antibody TU-20 and its scFv fragment

    Czech Academy of Sciences Publication Activity Database

    Kleinová, Veronika; Chaloupková, H.; Švecová, Helena; Fišer, Miroslav

    2010-01-01

    Roč. 286, č. 3 (2010), s. 847-851. ISSN 0236-5731 R&D Projects: GA AV ČR IBS1048301 Institutional research plan: CEZ:AV0Z10480505 Keywords : Monoclonal antibody TU-20 * ScFv TU-20 * Radiolabeling Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 0.777, year: 2010

  18. Non-linear dynamic modelling and design procedure of FV spring-dampers for base isolation

    OpenAIRE

    S. Sorace; Terenzi, G.

    2001-01-01

    Awarded Munro Prize 2001 jointly with “Non-linear dynamic design procedure of FV spring-dampers for base isolation - frame building applications”, (S. Sorace, G. Terenzi), Engineering Structures, Elsevier Science Ltd, Oxford, 23(12), pp. 1568-1576.

  19. An optimized Fermentation and Purification Process for a Recombinant Human Antibody ScFv-Fc Fragment Expressed in Pichia Pastoris%重组人小分子抗体ScFv-Fc发酵条件的优化及纯化

    Institute of Scientific and Technical Information of China (English)

    王丁丁; 苏曼曼; 胡丽莉; 袁丽颖; 颜炜群

    2011-01-01

    Objective: To obtain an optimized fermentation and purification process for high-level production of a recombinant human antibody ScFv-Fc fragment expression secreted in Pichia pastoris. Methods: The growth conditions of the transformant strain were optimized in 50 ml conical tubes including pH, methanol concentration and inducing time. The ScFv-Fc was purified using a two-step scheme: ammonium sulfate fractionation, protein A Sepharose. Results: ScFv-Fc production was found to increase with 0.5% (v/v) methanol concentration after 72 h induction. Protein production was also greatly affected by pH, resulting in higher yields at 5.2 pH value. The ScFv-Fc was purified more than 94% purity by using protein A Sepharose. Conclusions: The results provided a best process for expression and purification of functional recombinant human monoclonal antibody ScFv-Fc. It suggests a potential use of this antibody generating method by Pichia pastoris and indicates the potential of scFv-Fc fusion proteins as therapeutic candidates.%目的:研究重组人小分子抗体ScFv-Fc在毕赤酵母中分泌表达的最佳条件,以及ScFv-Fc的纯化方法.方法:分别从甲醇浓度、pH、诱导时间等方面对毕赤酵母重组菌株产生ScFv-Fc的发酵过程进行了优化;通过硫酸铵沉淀结合protein A亲和层析柱,对ScFv-Fc的纯化方法进行了研究.结果:确定ScFv-Fc在毕赤酵母中分泌表达的最佳条件为:在pH5.2的条件下,以0.5%甲醇诱导72 h.经过protein A亲和层析柱纯化后,ScFv-Fc纯度可达94%以上.结论:确定了ScFv-Fc在毕赤酵母中分泌表达的最佳条件以及纯化方法,为重组抗体分子诊断、治疗试剂的开发以及抗体的人源化奠定了物质基础.

  20. Construction and Biological Activity Detection of an Anti-Human Red Blood Cell Single Chain Fragment Variable-Pseudorabies Virus gE Bifunctional Fusion Protein%抗人红细胞单链抗体(ScFv)-伪狂犬病毒(PRV)gE蛋白双功能融合蛋白的构建及生物学活性检测

    Institute of Scientific and Technical Information of China (English)

    廖文军; 覃绍敏; 吴健敏; 王仰杰; 白安斌; 覃建飞

    2010-01-01

    利用特异性引物从重组质粒pMD-2E8ScFv和pMD-gE中PCR扩增出2E8基因(抗人红细胞H抗原单链抗体基因)和kgE基因(PRVgE主要抗原编码区),采用重叠延伸(SOE)PCR法拼接成融合的双功能分子2E8kgE,经BamH I和EcoR I双酶切,纯化后,克隆至BamH I和EcoR I双酶切的表达载体pET-Trx中,构建了原核表达载体pET-Trx-2E8kgE;将pET-Trx-2E8kgE转化至BL21plysS中,在IPTG诱导下表达具有双功能的融合蛋白,经过SDS-PAGE和Western-blot检测;结果表明,重组菌BL21plysS表达出约60.1 kD的目的蛋白,与猪伪狂犬标准阳性血清发生特异性反应.红细胞凝集试验证实,2E8kgE融合蛋白既能够与人红细胞结合,又能够与PRV抗体反应,具有双功能特性.

  1. 全价值链成本管控体系建设实践研究%Construction Practice Study of the System of Whole Value Chain Cost Management and Control

    Institute of Scientific and Technical Information of China (English)

    崔洁元; 韦英

    2015-01-01

    全价值链成本管控体系建设是以价值链管理和战略成本管理理念为指导,以作业成本管理为基础,全面收集、整理、分析和利用价值链上各环节的成本信息,以支持价值链成本管控体系的构建和优化。本文通过对全价值链成本管控进行理论阐述,在具体实践中运用精益管理理念,创新管理思维,理论联系实际,建设性地提出了推进全价值链成本管控体系建设的思路和具体实施方法,对构建企业全价值链成本管控体系有一定的借鉴意义。%Guided by value chain management and the theory of strategic cost management, and based on activity-based costing management, the whole value chain management system collets, sorts out, analyzes and makes use of the information on every value chain link to support the construction and optimization of value chain cost management system. The paper expounds the theory of whole value chain management and control by employing lean management concept, innovating management thinking, combining theory and practice, putting forward ideas and concrete measures to promote the construction of whole value chain management system, which may be reference to constructing whole value chain management system for enterprises.

  2. Production of in vivo biotinylated scFv specific to almond (Prunus dulcis) proteins by recombinant Pichia pastoris.

    Science.gov (United States)

    de la Cruz, Silvia; Alcocer, Marcos; Madrid, Raquel; García, Aina; Martín, Rosario; González, Isabel; García, Teresa

    2016-06-10

    The methylotropic yeast Pichia pastoris has demonstrated its suitability for large-scale production of recombinant proteins. As an eukaryotic organism P. pastoris presents a series of advantages at expression and processing of heterologous proteins when compared with Escherichia coli. In this work, P. pastoris has been used to express a scFv from a human synthetic library previously shown to bind almond proteins. In order to facilitate purification and post processing manipulations, the scFv was engineered with a C-terminal tag and biotinylated in vivo. After purification, biotinylated scFv were bound to avidin conjugated with HRP producing a multimeric scFv. The multimeric scFv showed to maintain their ability to recognize almond protein when assayed in ELISA, reaching a LOD of 470mgkg(-1). This study describes an easy method to produce large quantities of in vivo biotinylated scFv in P. pastoris. By substituting the enzyme or fluorochromes linked to avidin, it will be possible to generate a diverse number of multimeric scFv as probes to suit different analytical platforms in the detection of almond in food products. PMID:27085890

  3. Short-chain fluorescent tryptophan tags for on-line detection of functional recombinant proteins

    Directory of Open Access Journals (Sweden)

    Siepert Eva-Maria

    2012-09-01

    Full Text Available Abstract Background Conventional fluorescent proteins, such as GFP, its derivatives and flavin mononucleotide based fluorescent proteins (FbFPs are often used as fusion tags for detecting recombinant proteins during cultivation. These reporter tags are state-of-the-art; however, they have some drawbacks, which can make on-line monitoring challenging. It is discussed in the literature that the large molecular size of proteins of the GFP family may stress the host cell metabolism during production. In addition, fluorophore formation of GFP derivatives is oxygen-dependent resulting in a lag-time between expression and fluorescence detection and the maturation of the protein is suppressed under oxygen-limited conditions. On the contrary, FbFPs are also applicable in an oxygen-limited or even anaerobic environment but are still quite large (58% of the size of GFP. Results As an alternative to common fluorescent tags we developed five novel tags based on clustered tryptophan residues, called W-tags. They are only 5-11% of the size of GFP. Based on the property of tryptophan to fluoresce in absence of oxygen it is reasonable to assume that the functionality of our W-tags is also given under anaerobic conditions. We fused these W-tags to a recombinant protein model, the anti-CD30 receptor single-chain fragment variable antibody (scFv Ki-4(scFv and the anti-MucI single-chain fragment variable M12(scFv. During cultivation in Microtiter plates, the overall tryptophan fluorescence intensity of all cultures was measured on-line for monitoring product formation via the different W-tags. After correlation of the scattered light signal representing biomass concentration and tryptophan fluorescence for the uninduced cultures, the fluorescence originating from the biomass was subtracted from the overall tryptophan signal. The resulting signal, thus, represents the product fluorescence of the tagged and untagged antibody fragments. The product fluorescence signal

  4. Radioiodination and biodistribution of the monoclonal antibody TU-20 and its scFv fragment

    International Nuclear Information System (INIS)

    The ability of the monoclonal antibody TU-20 and its scFv fragment to bind specifically to the C-end of the class III β-tubulin makes these substances useful as potential diagnostics for neurodegenerative diseases-especially peripheral neuropathies. TU-20 and its scFv were labeled with 125I and 123I by chloramine-T (with radiochemical yield 75 and 50%, respectively). Radiochemical purity and stability was revealed by gel filtration (decrease to 80 and 50% in 2 months, respectively). Immunoreactivity of the labeled TU-20 was determined by ELISA - the range of the preserved immunoreactivity varies from 60 to 95% in accordance to the used radiolabeling process. RIA and affinity coupling analytic methods were specifically designed with focusing on specifics of the antibody and its fragment. The results of RIA differ in dependence on the type of the reaction vessel (glass or polystyrene) and the affinity coupling results depend on the experimental arrangement - in the batch or on the column. Fragmentation of the labeled antibody and its fragment was estimated by bis-tris gel electrophoresis followed by silver staining and autoradiography (over 95% of radioactivity bound in the substances). The antibody binding in tissue slices was studied in vitro by immunohistochemistry. The Purkinje cells were observed conjugated with the radiolabeled substances, either TU-20 or its ScFv fragment in the area of the cerebellum. In vivo biodistribution of 125I-TU-20, 125I-scFv TU-20, 123I-scFv TU-20 and Na125I was proceeded in normal mice (wild type C57B/6/J). Both biomolecules labeled by 123I were also proved in an imaging biodistribution study with use of the SPECT camera. Finally, a transgene population G93A1 Gur was used for comparative study to show the different behaviour of the substances in a normal mouse and in the modified organism with amyotrophic lateral sclerosis. The most part of differences is observed in the area of the muscles, rostal and caudal spinal cord. In summary

  5. PRODUCTION OF PHAGE-DISPLAYED ANTI-IDIOTYPIC ANTIBODY SINGLE CHAIN VARIABLE FRAGMENTS TO MG7 MONOCLONAL ANTIBODY DIRECTED AGAINST GASTRIC CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    何凤田; 聂勇战; 陈宝军; 乔太东; 韩者艺; 樊代明

    2002-01-01

    Objective. To generate phage-displayed anti-idiotypic antibody single chain variable fragments (anti -Id ScFv) to MG7 monoclonal antibody (McAb) directed against gastric carcinoma so as to lay a foundation for developing anti-Id ScFv vaccine of the cancer.Methods. Balb/c mice were immunized i. p. with MG7 McAb conjugated with keyhole limpet hemocyanin (KLH), and mRNA was isolated from the spleens of the immunized mice. Heavy and light chain (VH and VL)genes of antibody were amplified separately and assembled into ScFv genes with a linker DNA by PCR. The ScFv genes were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into competent E. coli TG1. The transformants were infected with M13KO7 helper phage to yield recombinant phages displaying ScFv on the tips of M13 phage. After 4 rounds of panning with MG7, the MG7-positive clones were selected by ELISA from the enriched phages. Thetypesoftheanti-IdScFvdisplayedontheselectedphagecloneswerepreliminarily identified by competition ELISA.Results. The VH, VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. Twenty-four MG7-positive clones were selected from 60 enriched phage clones, among which 5 displayed β or γtype anti-Id ScFv.Conclsion. The anti-Id ScFv to MG7 McAb can be successfully selected by recombinant phage antibody technique, which paves a way for the study of prevention and cure of gastric carcinoma by using anti-Id ScFv.

  6. Expression and Functional Properties of an Anti-Triazophos High-Affinity Single-Chain Variable Fragment Antibody with Specific Lambda Light Chain

    Directory of Open Access Journals (Sweden)

    Rui Liu

    2016-06-01

    Full Text Available Triazophos is a widely used organophosphorous insecticide that has potentially adverse effects to organisms. In the present study, a high-affinity single-chain variable fragment (scFv antibody with specific lambda light chain was developed for residue monitoring. First, the specific variable regions were correctly amplified from a hybridoma cell line 8C10 that secreted monoclonal antibody (mAb against triazophos. The regions were then assembled as scFv via splicing by overlap extension polymerase chain reaction. Subsequently, the recombinant anti-triazophos scFv-8C10 was successfully expressed in Escherichia coli strain HB2151 in soluble form, purified through immobilized metal ion affinity chromatography, and verified via Western blot and peptide mass fingerprinting analyses. Afterward, an indirect competitive enzyme-linked immunosorbent assay was established based on the purified anti-triazophos scFv-8C10 antibody. The assay exhibited properties similar to those based on the parent mAb, with a high sensitivity (IC50 of 1.73 ng/mL to triazophos and no cross reaction for other organophosphorus pesticides; it was reliable in detecting triazophos residues in spiked water samples. Moreover, kinetic measurement using a surface plasmon resonance biosensor indicated that the purified scFv-8C10 antibody had a high affinity of 1.8 × 10−10 M and exhibited good binding stability. Results indicated that the recombinant high-affinity scFv-8C10 antibody was an effective detection material that would be promising for monitoring triazophos residues in environment samples.

  7. Round table on the Supply Chain for NPPs construction: Localization - Daya Bay Experience; REEL Handling and Lifting Systems, More than 60 years expertise in lifting and handling equipment in production process

    International Nuclear Information System (INIS)

    The second day afternoon began with the round table on the Supply Chain for NPPs construction with Philippe Frantz, President of REEL, Marc Lachaise, Head of procurement of NNB at EDF Energy, and Steven Lau, First Deputy General Manager of DNMC. Philippe Frantz started to present the activities and the contribution of REEL in the construction of NPPs as a main supplier of handling system. Then, Marc Lachaise took the lead to present Hinkley Point C Project, the Values of NNB and the key role of the supply chain in this Project. Steven Lau went on to describe the link of the supply chain with the operating of NPPs and explained the cooperation between EDF and CGNPC in order to secure the supply of equipment. Following their presentation, they started the open discussion with the audience by explaining their strategy to make or to buy and the link of this strategy to their core business. They also highlighted the new relations and the new partnership between supplier and customer. They insisted on the necessity to invest on supply chain and to have a strong Nuclear Safety Culture in the supply chain

  8. 基于电子商务技术的供应链系统构建分析%The Analysis of the Construction of Supply Chain System Based on E-commerce Technology

    Institute of Scientific and Technical Information of China (English)

    张慧

    2013-01-01

    Based on the research on the relationship between supply chain system and e-commerce the necessity of constructing the supply chain for e-commerce is analyzed and summered up. Consequently, the strategies for constructing a supply chain for e-commerce are proposed starting from the characteristics of the supply chain system for e-commerce environment, so as to further improve the suitability of supply chain system in the development of e-commerce.%本文在研究供应链系统和电子商务相互关系的基础上,分析得出构建电子商务供应链的必要性,从而从电子商务供应链的特点出发得出如何构建一个电子商务供应链的策略,从而进一步提高供应链系统在电子商务发展中的适应性。

  9. DEVELOPMENT AND COMMERCIAL APPLICATION OF FV-10 CATALYST FOR PARAFFIN WAX HYDROFINING%FV-10石蜡加氢精制催化剂的开发及工业应用

    Institute of Scientific and Technical Information of China (English)

    王士新; 袁平飞; 李殿昭

    2007-01-01

    介绍了FV-10石蜡加氢精制催化剂石蜡及微晶蜡加氢试验情况,表明FV-10催化剂具有较高活性和稳定性,其性能优于FV-1催化剂和481-2B催化剂,可用于食品级石蜡和白色食品级微晶蜡生产.FV-10催化剂已在三套工业装置上应用,装置运转平稳,产品质量合格,达到预期效果.

  10. Construction of copper-based coordination polymers with 1D chain, 2D plane and wavy networks: Syntheses, structures, thermal behaviors and photoluminescence properties

    Indian Academy of Sciences (India)

    Jianghua Li; Chi Zhang

    2015-11-01

    Three Cu-based coordination polymers (CPs), including [CuII ( -1 -NCS)2 (O-1 -DMF)2 (2 -3,3’-bptz)] (1), [CuI (1,3-2-NCS)(2-3,3’-bptz)] (2) and [(CuI (1,3-μ2- NCS))(2-4,4’-bptz)] (3) (DMF = , -dimethyl formamide, 3,3’-bptz = 3,6-bis(3-pyridyl)tetrazine and 4,4’-bptz = 3,6-bis(4-pyridyl)tetrazine) have been successfully constructed by solution diffusion reactions by using Cu(NO3)2 ·3H2O or CuNCS and KNCS with 3,3’-bptz / 4,4’-bptz ligands, respectively. The resulting crystalline materials have been characterized by the single-crystal X-ray diffraction analyses, elemental analyses, FT-IR spectra, thermogravimetric analyses and powder X-ray diffraction (PXRD). Single crystal X-ray analyses revealed that CP 1 is organized in one-dimensional (1D) chain in which the Cu(II) ions are coordinated by 1 -NCS− anions and 1-DMF molecules, and linked by 2-3,3’-bptz bridging ligands. CPs ,2 and 3 are structural isomers. CP 2 exhibits two-dimensional (2D) (4,4)-plane-like network in which Cu(I) ions are linked by 2-NCS − and 2-3,3’-bptz ligands. In CP 3, Cu(I) ions are connected by 2 -NCS − and 2-4,4’-bptz ligands to form 2D saw-tooth wavy network. In addition, the photoluminescence properties of CPs 1-3 were also investigated in the solid state at room temperature.

  11. Interstitial loop growth in electron irradiated vacuum-remelted FV448

    International Nuclear Information System (INIS)

    An investigation of the resistance to void swelling of the ferritic steel FV448 has been made in electron irradiation experiments in a High Voltage Electron Microscope. Overaged samples of vacuum-remelted FV448 showed zero swelling after irradiation to doses of up to 30 dpa, in the temperature range 400 to 5500C. The resistance to void swelling was due to the absence of mutual interaction between interstitial loops. Impurity segregation to the loops, and precipitation, prevented continuous dislocation climb, thus removing biased sinks from the system at low doses. An interstitial-impurity binding energy of 0.5 eV was measured, suggesting that the impurity responsible was carbon or nitrogen. Some effects attributable to the heating of thin foils in the HVEM environment were detected. (author)

  12. A Successful Mother and Neonate Outcome for a Woman with Essential Thrombocytosis and FV Leiden Heterozygosity

    Directory of Open Access Journals (Sweden)

    Marianna Politou

    2016-01-01

    Full Text Available Essential thrombocytosis (ET and FV Leiden heterozygosity represent an acquired and hereditable hypercoagulable state, respectively. An uncommon case of coexistence of ET and FV Leiden heterozygosity in a 36-year-old pregnant woman and her successful pregnancy outcome is described. She was considered to be at high risk of thrombosis during her pregnancy and she was treated with both prophylactic dose of LMWH and aspirin daily throughout her pregnancy and for a 6-week period postpartum. The efficacy of the anticoagulation treatment was monitored in various time points not only by measuring anti-Xa levels and D-Dimers but also with new coagulation methods such as rotation thromboelastometry and multiplate. Global assessment of coagulation using additional newer laboratory tests might prove useful in monitoring coagulation pregnancies at high risk for thrombosis.

  13. Non-linear dynamic design procedure of FV spring-dampers for base isolation - Frame building applications

    OpenAIRE

    S. Sorace; Terenzi G.

    2001-01-01

    Awarded Munro Prize 2001 jointly with “Non-linear dynamic modelling and design procedure of FV spring-dampers for base isolation”, (S. Sorace, G. Terenzi). Engineering Structures, Elsevier Science Ltd, Oxford, 23(12), pp. 1556-1567.

  14. A bacterial signal peptidase enhances processing of a recombinant single chain antibody fragment in insect cells

    NARCIS (Netherlands)

    Ailor, E; Pathmanathan, J; Jongbloed, JDH; Betenbaugh, MJ

    1999-01-01

    The production of an antibody single chain fragment (scFv) in insect cells was accompanied by the formation of an insoluble intracellular precursor even with the inclusion of the bee melittin signal peptide. The presence of the precursor polypeptide suggests a limitation in the processing of the sig

  15. Single Chain Variable Fragments Produced in Escherichia coli against Heat-Labile and Heat-Stable Toxins from Enterotoxigenic E. coli.

    Directory of Open Access Journals (Sweden)

    Christiane Y Ozaki

    Full Text Available Diarrhea is a prevalent pathological condition frequently associated to the colonization of the small intestine by enterotoxigenic Escherichia coli (ETEC strains, known to be endemic in developing countries. These strains can produce two enterotoxins associated with the manifestation of clinical symptoms that can be used to detect these pathogens. Although several detection tests have been developed, minimally equipped laboratories are still in need of simple and cost-effective methods. With the aim to contribute to the development of such diagnostic approaches, we describe here two mouse hybridoma-derived single chain fragment variable (scFv that were produced in E. coli against enterotoxins of ETEC strains.Recombinant scFv were developed against ETEC heat-labile toxin (LT and heat-stable toxin (ST, from previously isolated hybridoma clones. This work reports their design, construction, molecular and functional characterization against LT and ST toxins. Both antibody fragments were able to recognize the cell-interacting toxins by immunofluorescence, the purified toxins by ELISA and also LT-, ST- and LT/ST-producing ETEC strains.The developed recombinant scFvs against LT and ST constitute promising starting point for simple and cost-effective ETEC diagnosis.

  16. On the Application of Value Chain Theory in Construction Engineering Enterprise Cost Management%浅析价值链理论在建筑工程企业成本管理的应用

    Institute of Scientific and Technical Information of China (English)

    陆乃炎; 韩吉鹏

    2013-01-01

    On the basis of value chain theory, this paper points out the common defects in the cost management of construction engineering in China and then analyzes the relationship between value chain and cost control. Finally, the outstanding contribution of value chain theory to constriction engineering cost management is analyzed.%本文在介绍价值链理论的基础上,提出了我国建筑工程成本管理普遍存在的缺陷,接着对价值链与成本控制的关系进行分析,最后分析价值链在建筑工程成本管理中的突出贡献。

  17. Production and characterization of a biotinylated single-chain variable fragment antibody for detection of parathion-methyl.

    Science.gov (United States)

    Wang, Huimin; Zhao, Fengchun; Han, Xiao; Yang, Zhengyou

    2016-10-01

    In this article, we reported the development of a biotinylated single-chain variable fragment (scFv) antibody based indirect competitive enzyme-linked immunosorbent assay (IC-ELISA) for parathion-methyl (PM) detection. Firstly, a phage display library was generated using a pre-immunized BALB/C mouse against a specific hapten of PM. After four rounds of panning, the scFv gene fragments were transferred into a secreted expression vector. Then, the scFv antibodies were secreted expressed and screened by IC-ELISA against PM. The selected scFv antibody was fused with a biotin acceptor domain (BAD) and inserted into pET-28a(+) vector for high-level expression in Escherichia coli BL2 (DE3). After optimizing expression conditions, the scFv-BAD antibody was expressed as a soluble protein and biotinylated in vitro by the E. coli biotin ligase (BirA). Subsequently, the biotinylated scFv-BAD antibody was purified with a high yield of 59.2 ± 3.7 mg/L of culture, and was characterized by SDS-PAGE and western blotting. Finally, based on the biotinylated scFv-BAD, a sensitive IC-ELISA for detection of PM was developed, and the 50% inhibition value (IC50) of PM was determined as 14.5 ng/mL, with a limit of detection (LOD, IC10) of 0.9 ng/mL. Cross-reactivity (CR) studies revealed that the scFv antibody showed desirable specificity for PM. PMID:27181246

  18. Affinity improvement by fine tuning of single-chain variable fragment against aflatoxin B1.

    Science.gov (United States)

    Min, Won-Ki; Na, Kang-In; Yoon, Jung-Hyun; Heo, Yoon-Jee; Lee, Daesang; Kim, Sung-Gun; Seo, Jin-Ho

    2016-10-15

    Aflatoxin B1 (AFB1) produced in Aspergillus flavus is a major hepatocarcinogen found in foods and feed. For effective immunological detection of AFB1 at low concentrations, the development of high affinity antibody for AFB1 is required. Previously, an affinity-maturated single-chain variable fragment containing 6 mutations (scFv-M37) was isolated from an artificial mutagenic library, which showed a 9-fold higher affinity than its wild type scFv. In this study, the effect of the 6 mutated residues on the affinity improvement was characterized using surface plasmon resonance analysis, which identified a deleterious mutation (VH-A110T) located on a framework region of the scFv-M37. The back mutation of VH-A110T resulted in a 3.2-fold affinity improvement, which was attributed to decrease of dissociation rate constant (kd) in interaction between AFB1 and the back mutant scFv. The biophysical analyses using circular dichroism and gel filtration revealed that the back mutation of VH-A110T caused a subtle conformational change of the scFv toward tighter binding to AFB1. PMID:27173568

  19. PURE mRNA display for in vitro selection of single-chain antibodies.

    Science.gov (United States)

    Nagumo, Yu; Fujiwara, Kei; Horisawa, Kenichi; Yanagawa, Hiroshi; Doi, Nobuhide

    2016-05-01

    mRNA display is a method to form a covalent linkage between a cell-free synthesized protein (phenotype) and its encoding mRNA (genotype) through puromycin for in vitro selection of proteins. Although a wheat germ cell-free translation system has been previously used in our mRNA display system, a protein synthesis using recombinant elements (PURE) system is a more attractive approach because it contains no endogenous nucleases and proteases and is optimized for folding of antibodies with disulphide bonds. However, when we used the PURE system for mRNA display of single-chain Fv (scFv) antibodies, the formation efficiency of the mRNA-protein conjugates was quite low. To establish an efficient platform for the PURE mRNA display of scFv, we performed affinity selection of a library of scFv antibodies with a C-terminal random sequence and obtained C-terminal sequences that increased the formation of mRNA-protein conjugates. We also identified unexpected common substitution mutations around the start codon of scFv antibodies, which were inferred to destabilize the mRNA secondary structure. This destabilization causes an increase in protein expression and the efficiency of the formation of mRNA-protein conjugates. We believe these improvements should make the PURE mRNA display more efficient for selecting antibodies for diagnostic and therapeutic applications. PMID:26711234

  20. Relacionamento interorganizacional na cadeia de suprimentos: um estudo de caso na indústria da construção civil The interorganizational relationship in the supply chain: a case study in the civil construction industry

    Directory of Open Access Journals (Sweden)

    Renata Albergaria de Mello Bandeira

    2009-01-01

    Full Text Available O escopo deste trabalho consiste em promover uma discussão sobre os mecanismos de coordenação das relações interorganizacionais - poder e cooperação - na cadeia de suprimentos da indústria da construção civil. O poder, como um construto de relações interorganizacionais, tem recebido um tratamento irregular e conflitante por parte dos analistas. No entanto, esta abordagem ignora relações existentes que são muito apropriadas para certos contextos do relacionamento interfirmas. Logo, foi desenvolvido um estudo de caso em um elo da indústria da construção civil com o intuito de identificar suas relações como sendo baseadas em dominação ou cooperação. São analisadas, ainda, as implicações dos resultados da pesquisa na gestão da cadeia de suprimentos da construção civil.The article discusses the relationship in the supplier chain in the civil construction industry. The main objective is to define if it is a cooperative relation among the intervenients or if the most powerful dictates the rules. Power as an interorganizational construct has been observed by analysts with an irregular and conflictive point of view. However, these analysts ignore completely existing relationships that are very appropriate in some existing relationships among enterprises. Due to that, it was verified het type of relationship developed by two intervenients in one step of the supply chain of a civil construction company to determine if it is cooperative or if, otherwise, the main actor uses its power to lead the chain. The researchers also analyze what are the implications of this study in the civil construction industry's supply management.

  1. Affinity maturation of anti-TNF-alpha scFv with somatic hypermutation in non-B cells.

    Science.gov (United States)

    Chen, Shaopeng; Qiu, Junkang; Chen, Chuan; Liu, Chunchun; Liu, Yuheng; An, Lili; Jia, Junying; Tang, Jie; Wu, Lijun; Hang, Haiying

    2012-06-01

    Activation-induced cytidine deaminase (AID) is required for the generation of antibody diversity through initiating both somatic hypermutation (SHM) and class switch recombination. A few research groups have successfully used the feature of AID for generating mutant libraries in directed evolution of target proteins in B cells in vitro. B cells, cultured in suspension, are not convenient for transfection and cloning. In this study, we established an AID-based mutant accumulation and sorting system in adherent human cells. Mouse AID gene was first transfected into the human non-small cell lung carcinoma H1299 cells, and a stable cell clone (H1299-AID) was selected. Afterwards, anti-hTNF-α scFv (ATscFv) was transfected into H1299-AID cells and ATscFv was displayed on the surface of H1299-AID cells. By 4-round amplification/flow cytometric sorting for cells with the highest affinities to hTNF-alpha, two ATscFv mutant gene clones were isolated. Compared with the wild type ATscFv, the two mutants were much more efficient in neutralizing cytotoxicity of hTNF-alpha. The results indicate that directed evolution by somatic hypermutation can be carried out in adherent non-B cells, which makes directed evolution in mammalian cells easier and more efficient. PMID:22467272

  2. scFv from Antibody That Mimics gp43 Modulates the Cellular and Humoral Immune Responses during Experimental Paracoccidioidomycosis.

    Directory of Open Access Journals (Sweden)

    Grasielle Pereira Jannuzzi

    Full Text Available Paracoccidioidomycosis (PCM, caused by Paracoccidioides species is a prevalent systemic and progressive mycosis that occurs in Latin America. It is caused by Paracoccidioides species. Immunization with dendritic cells transfected with a plasmid encoding the scFv (pMAC/PS-scFv that mimics the main antigen of P. brasiliensis (gp43 confers protection in experimental PCM. DCs link innate and adaptive immunity by recognizing invading pathogens and selecting the type of effector T cell to mediate the immune response. Here, we showed that DC-pMAC/PS-scFv induces the activation of CD4+ and CD8+ T cells. Moreover, our results demonstrated that BALB/c mice infected with P. brasiliensis and treated with DC-pMAC/PS-scFv showed the induction of specific IgG production against gp43 and IFN-γ, IL-12 and IL-4 cytokines. Analysis of regional lymph nodes revealed increases in the expression of clec7a, myd88, tlr2, gata3 and tbx21, which are involved in the immune response. Taken together, our results indicate that the scFv modulates the humoral and cellular immune responses and presents epitopes to CD4+ and CD8+ T cells.

  3. Putting Markov Chains Back into Markov Chain Monte Carlo

    OpenAIRE

    Barker, Richard J.; Schofield, Matthew R.

    2007-01-01

    Markov chain theory plays an important role in statistical inference both in the formulation of models for data and in the construction of efficient algorithms for inference. The use of Markov chains in modeling data has a long history, however the use of Markov chain theory in developing algorithms for statistical inference has only become popular recently. Using mark-recapture models as an illustration, we show how Markov chains can be used for developing demographic models and also ...

  4. Prediction of FV520B Steel Flow Stresses at High Temperature and Strain Rates

    Science.gov (United States)

    Han, Xiaolan; Zhao, Shengdun; Zhang, Chenyang; Fan, Shuqin; Xu, Fan

    2015-10-01

    In order to develop reliable constitutive equations for the simulation, the hot deformation behavior of FV520B steel was investigated through isothermal compression tests in a wide range of temperatures from 900 °C to 1100 °C at an interval of 50 °C and strain rate from 0.01 to 10 s-1 on Gleeble-1500D simulator. The effects of temperature and strain rate on deformation behavior were represented by Zener-Holloman parameter in an exponent-type equation of Arrhenius constitutive. The influence of strain was incorporated in the constitutive analysis by material constants expressed as a polynomial function of strain. The constitutive equation (considering the compensation of strain) could precisely predict the flow stress only at strain rate 0.01 s-1 except at the temperatures of 900 °C and 1000 °C, whereas the flow stress predicted by a modified equation (incorporating both the strain and strain rate) demonstrated a well agreement with the experimental data throughout the entire range of temperatures and strain rates. Correlation coefficient (R) of 0.988 and average absolute relative error (AARE) of 5.7% verified the validity of developed equation from statistical analysis, which further confirmed that the modified constitutive equation could accurately predict the flow stress of FV520B steel.

  5. 建筑施工企业供应链管理现状及存在的问题%Supply-chain management status and existing problems of building construction enterprise

    Institute of Scientific and Technical Information of China (English)

    史国红

    2011-01-01

    概括了我国建筑施工企业供应链管理现状,根据项目流程总结了目前建筑供应链管理亟待解决的问题,并剖析了其原因,最后提出先进管理手段,以实现企业与供应商之间的互通。%This thesis outlines supply-chain management status of building construction enterprise,summarizes urgent problems needing solving in temporary building supply-chain management according to project procedure,and analyzes its causes.In the end,it proposes advanced management means,with a view to realize mutual connection between enterprise and supplier.

  6. Supply chain management in product development.

    OpenAIRE

    Tzortzopoulos, Patricia; Kagioglou, Mike; Cooper, Rachel; Dyson, E

    2009-01-01

    Mounting emphasis on construction supply chain management (CSCM) is due to both global sourcing of materials and a shortage of labor. These factors force increasing amounts of value-added work to be conducted off-site deep in the supply chain. Construction Supply Chain Management Handbookcompiles in one comprehensive source an overview of the diverse research and examples of construction supply chain practice around the world.Reflecting the emergence of CSCM as an important area of multi-nati...

  7. Engineering of FRT-lacZ fusion constructs: induction of the Pseudomonas aeruginosa fadAB1 operon by medium and long chain-length fatty acids

    OpenAIRE

    Son, Mike S.; Nguyen, David T.; Kang, Yun (Kenneth); Hoang, Tung T.

    2008-01-01

    Without prior knowledge of the promoters of various genes in bacteria, it can be difficult to study gene regulation using reporter-gene fusions. Regulation studies of promoters are ideal at their native locus, which do not require prior knowledge of promoter regions. Based on a previous study with FRT-lacZ-KmR constructs, we constructed two novel FRT-lacZ-GmR plasmids. This allows easy engineering of P. aeruginosa reporter-gene fusions, post-mutant construction with the Flp-FRT system. We dem...

  8. 影视旅游价值链构建及发展策略研究%Construction of Film and TV Tourism Value Chain and Development Strategy

    Institute of Scientific and Technical Information of China (English)

    曹菲

    2013-01-01

      This paper uses the value chain theory of professor Michael E. Porter, sets up the value chain of film and TV tourism, designs the activities content of each link from main body activities and auxiliary activities, and analyzes the development of film and TV tourism from the angle of node activity strategy.%  本文运用Michael E. Porter教授提出的价值链理论,建立影视旅游价值链,从主体活动和辅助活动两个角度设计各个环节的活动内容,并从节点活动角度分析了发展影视旅游的策略。

  9. Quantity Flexibility Contract Based on Construction Supply Chain Management%基于建筑供应链管理的工程采购数量弹性契约研究

    Institute of Scientific and Technical Information of China (English)

    査京民; 黄雷

    2013-01-01

    Adoption of integrated supply chain management is the key to reduce cost in construction industry. Aiming at the disadvantages of low degree cooperation,inefficient purchasing processes,and poor adaptability to demand uncertainty,the traditional purchasing model and the purchasing model under construction supply chain management environment were compared. The quantity flexibility purchasing contract model was set up based on contract theory,which proves that the contract model can effectively cope with demand uncertainty,enable contractor to flexibly carry out certain purchasing activity and further coordinate the construction supply chain.%建筑业采用集成式的供应链管理是降低建设工程成本的重要方法。为解决传统工程采购模式存在的合作度低、采购效率低下、对不确定性的应变能力差等缺陷。对比了建筑供应链管理环境下与传统工程采购模式的特点,基于供应链契约理论,构建了承包商与供应商之间的数量弹性采购契约模型,并证明了该契约模式可以有效应对工程采购中需求的不确定性。结论表明,建筑供应链管理下的数量弹性采购契约模式可以使承包商灵活地进行特定的采购活动,进而使建筑供应链上下游企业在一定程度上达到协调。

  10. 基于价值链管理的内部控制构建思想%Constructing Thoughts of Internal Control Based on Value Chain Management

    Institute of Scientific and Technical Information of China (English)

    张其镇

    2011-01-01

    Development and application of information technology and value chain theory provide a good opportunity to improve the management level of enterprise and gain a competitive advantage. This paper, combining with the value chain theory, analyzes the key steps to build the internal control which can adapt to the management of value chain under the guidance of the internal control structure. This paper .argues that internal control should be value-added activities which take getting a competitive advantage as the strategic goal, take theall kinds of "flow" in business as the main line of control and constantly self-perfect.%信息技术与价值链理论的发展与应用,为提高企业管理水平,获得竞争优势提供了一个良好的契机.本文结合价值链理论,分析了在内部控制结构指导下构建一个能适应价值链管理的内部控制的关键步骤.本文认为,内部控制应是一个以获取竞争优势为战略目标,以企业内各种"流"为控制主线并能不断自我完善的增值活动.

  11. Based on JMI Model the Study in Implementation Strategy of Construction Supply Chain%基于JMI模式的建筑供应链实施策略研究

    Institute of Scientific and Technical Information of China (English)

    刘新欣; 王红春

    2014-01-01

    工程建设项目消耗的物资数量大、品种多,能否合理安排物资储备,直接关系到建设项目的总进度和总成本。随着建筑供应链思想的不断深化,结合工程建设自身的特点,文中提出联合库存管理模式,明确应用条件及实施策略,可以有效地降低施工企业的总库存成本,加快企业的资金周转速率,实现供应商与建筑企业之间的战略共赢,最终提高建筑企业的综合竞争力。%The number of construction projects goods consumed in large and variety,set up arrangements for material reserves sensibly,directly related to the overall progress and the total cost of the construction project. With the continue deepening of the construction supply chain ideology,combined with the characteristics of construction,the paper proposes the joint management inventory,nail down application conditions and implementation strategy,reduce the total inventory cost of construction enterprises effectively,accelerate the capital turnover rate of enterprises,realize win-win strategy between providers and construction companies,at last,improve the overall competitiveness of construction enterprises.

  12. Value Chain Theory in Agent Construction Project Investment Control:From Construction Agency Perspective%价值链理论在代建项目投资控制中的应用研究--基于代建企业视角

    Institute of Scientific and Technical Information of China (English)

    刘逸凡; 刘万琳

    2013-01-01

    From the view of the Construction Agency,this paper used the value chain analysis model to study investment control within the whole process of the Agent Construction Project,proposed to establish a internal value chain which involves two key segments (the optimization of the design choices in the design phase and the control of design changes in construction phase) of the Construction Agency, thus successfully control the project investment. Through the value chain support systems,sort out all the value activities within theses two key segments,draw conclusions of effective measures that control the project investment,thereby improve the management performance of Construction Agency and enable them to gain a competitive advantage during the bidding process that held by government.%  本文从代建企业的角度出发,利用价值链理论分析模型对代建项目全过程的投资控制进行研究,提出以设计阶段设计方案的优化选择和施工阶段设计变更的控制作为企业内部价值链两大关键环节,从而有效地对代建项目进行投资控制。通过价值链中的支持系统,梳理关键环节中的价值活动,得出代建项目投资控制的有效措施,进而提高代建企业的管理绩效,在政府委托招标过程中获得竞争优势。

  13. Generation of a Highly Reactive Chicken-Derived Single-Chain Variable Fragment against Fusarium verticillioides by Phage Display

    Directory of Open Access Journals (Sweden)

    Zu-Quan Hu

    2012-06-01

    Full Text Available Fusarium verticillioides is the primary causal agent of Fusarium ear and kernel rot in maize, producing fumonisin mycotoxins that are toxic to humans and domestic animals. Rapid detection and monitoring of fumonisin-producing fungi are pivotally important for the prevention of mycotoxins from entering into food/feed products. Chicken-derived single-chain variable fragments (scFvs against cell wall-bound proteins from F. verticillioides were isolated from an immunocompetent phage display library. Comparative phage enzyme-linked immunosorbant assays (ELISAs and sequencing analyses identified four different scFv antibodies with high sensitivity. Soluble antibody ELISAs identified two highly sensitive scFv antibodies, FvCA3 and FvCA4, with the latter being slightly more sensitive. Three-dimensional modeling revealed that the FvCA4 may hold a better overall structure with CDRH3, CDRL1 and CDRL3 centered in the core region of antibody surface compared with that of other scFvs. Immunofluorescence labeling revealed that the binding of FvCA4 antibody was localized to the cell walls of conidiospores and hyphae of F. verticillioides, confirming the specificity of this antibody for a surface target. This scFv antibody was able to detect the fungal mycelium as low as 10−2 μg/mL and contaminating mycelium at a quantity of 10−2 mg/g maize. This is the first report that scFv antibodies derived from phage display have a wide application for rapid and accurate detection and monitoring of fumonisin-producing pathogens in agricultural samples.

  14. Insight into earthquake sequencing: analysis and interpretation of time-series constructed from the directed graph of the Markov chain model

    Science.gov (United States)

    Cavers, M. S.; Vasudevan, K.

    2015-02-01

    Directed graph representation of a Markov chain model to study global earthquake sequencing leads to a time-series of state-to-state transition probabilities that includes the spatio-temporally linked recurrent events in the record-breaking sense. A state refers to a configuration comprised of zones with either the occurrence or non-occurrence of an earthquake in each zone in a pre-determined time interval. Since the time-series is derived from non-linear and non-stationary earthquake sequencing, we use known analysis methods to glean new information. We apply decomposition procedures such as ensemble empirical mode decomposition (EEMD) to study the state-to-state fluctuations in each of the intrinsic mode functions. We subject the intrinsic mode functions, the orthogonal basis set derived from the time-series using the EEMD, to a detailed analysis to draw information-content of the time-series. Also, we investigate the influence of random-noise on the data-driven state-to-state transition probabilities. We consider a second aspect of earthquake sequencing that is closely tied to its time-correlative behavior. Here, we extend the Fano factor and Allan factor analysis to the time-series of state-to state transition frequencies of a Markov chain. Our results support not only the usefulness the intrinsic mode functions in understanding the time-series but also the presence of power-law behaviour exemplified by the Fano factor and the Allan factor.

  15. A Novel Three-dimensional Inorganic Open-framework Constructed from [Mo4O16]n8n- Chains Linked through Ag+

    Institute of Scientific and Technical Information of China (English)

    GUO Hong-Xu; LIU Shi-Xiong

    2005-01-01

    A novel 3-D inorganic bimetallic compound Ag2Mo2O7 has been hydrothermally synthesized and structurally characterized by X-ray diffraction. The compound crystallizes in mono- clinic, space group P21/c with a = 6.1274(3), b = 13.1836(6), c = 7.8780(3)(A), β = 110.673(3)(A), V = 595.42(5)(A)3, Z = 4, Mr = 519.62, Dc = 5.797 g/cm3, F(000) = 936, μ(MoKα) = 10.579 mm-1, F(000) = 936, the final R = 0.0204 and wR = 0.0477 for 1268 observed reflections with I > 2σ(I). In the title compound, the [Mo4O16]n8n- chains are linked through O-Ag-O bridging bonds to form a 3D network. It represents an unprecedented linking mode between the isopolyoxomolybdate chain and the transition metal oxide cluster. IR, TGA and fluorescence properties of the title compound are also studied.

  16. 生产性服务业:构建中国制造业国家价值链的关键%Producer Services: the Key to Constructing the National Value Chain of Chinese Manufacturing

    Institute of Scientific and Technical Information of China (English)

    贾根良; 刘书瀚

    2012-01-01

    生产性服务业在全球价值链的系统整合中处于关键性地位,拥有强大的研发能力并掌控整个价值链是生产性服务业的核心功能。依托巨大的国内市场,以生产性服务业为龙头,构建独立自主的制造业国家价值链,是中国制造业转型升级的根本措施。在中国制造业国家价值链的构建中,生产性服务业主要通过四种途径发挥着核心作用:中国制造业的领导型企业作为系统整合者主要执行的是包括研发在内的生产性服务功能,这是成长为中国跨国公司的基本途径;专业化市场交易平台承担着国内市场高度组织化并获取国际定价权的重要职能;国家价值链的国外布局是构建中国制造业全球价值链的初步尝试;电子商务革命和连锁专卖体系等新型生产性服务业的兴起,不仅是中国制造业企业打破营销商垄断国内终端销售渠道的有力途径,也是其突破跨国公司垄断发达国家终端销售市场的历史机遇。%Producer services, whose core functions are possessing strong R&D capabilities and controlling the whole value chain, are in a key position in the system integration of global value chain. The fundamental measure for the transforming and upgrading of Chinese manufacturing is to construct independent national manufacturing value chain with producer services as the leader, and relies on huge domestic market. During the construction of the national manufacturing value chain, producer services play a central role mainly in four ways: As system integrators, China' s leading manufacturing enterprises mainly perform the productive service functions which include research and development, and this is the basic way for these enterprises to grow into multinational corporations; Specialized market trading platform undertakes vital responsibilities to build a highly organized domestic market and gain international pricing power; The layout of

  17. Value Analysis of Construction Projects under the Lean Supply Chain%基于精益供应链的工程建设项目价值分析

    Institute of Scientific and Technical Information of China (English)

    谢秋玲; 叶青

    2012-01-01

    Combining with the lean supply chain theory and taking the value as the key of solution and decision-making, value analysis of construction project is studied, and the application process of value analysis is also illustrated by case study. The value of each solution is comprehensively evaluated by fuzzy quality function deployment, and compared with each other to find the optimal one. The value analysis of construction projects under the lean supply chain is an effective decision-making method.%结合精益供应链理论,以价值作为方案决策要点,对工程建设项目价值分析进行研究,通过案例说明价值分析在工程建设项目中的应用流程.采用模糊质量功能展开法综合评价各方案的价值,并进行对比分析,找出价值最优方案.基于精益供应链的工程建设项目价值分析是一种行之有效的方案决策方法.

  18. Comparative response of platelet fV and plasma fV to activated protein C and relevance to a model of acute traumatic coagulopathy.

    Directory of Open Access Journals (Sweden)

    James E Campbell

    Full Text Available BACKGROUND: Acute traumatic coagulopathy (ATC has been linked to an increase in activated protein C (aPC from 40 pM in healthy individuals to 175 pM. aPC exerts its activity primarily through cleavage of active coagulation factor Va (fVa. Platelets reportedly possess fVa which is more resistant to aPC cleavage than plasma fVa; this work examines the hypothesis that normal platelets are sufficient to maintain coagulation in the presence of elevated aPC. METHODS: Coagulation responses of normal plasma, fV deficient plasma (fVdp, and isolated normal platelets in fVdp were conducted: prothrombin (PT tests, turbidimetry, and thromboelastography (TEG, including the dose response of aPC on the samples. RESULTS: PT and turbidimetric assays demonstrate that normal plasma is resistant to aPC at doses much higher than those found in ATC. Additionally, an average physiological number of washed normal platelets (200,000 platelets/mm3 was sufficient to eliminate the anti-coagulant effects of aPC up to 10 nM, nearly two orders of magnitude above the ATC concentration and even the steady-state pharmacological concentration of human recombinant aPC, as measured by TEG. aPC also demonstrated no significant effect on clot lysis in normal plasma samples with or without platelets. CONCLUSIONS: Although platelet fVa shows slightly superior resistance to aPC's effects compared to plasma fVa in static models, neither fVa is sufficiently cleaved in simulations of ATC or pharmacologically-delivered aPC to diminish coagulation parameters. aPC is likely a correlative indicator of ATC or may play a cooperative role with other activity altering products generated in ATC.

  19. 基于精益价值链理论的建筑业企业精益建造能力评价研究%Evaluation of Construction Enterprise’s Lean Construction Ability Based on the Theory of Lean Value Chain

    Institute of Scientific and Technical Information of China (English)

    陈礼靖; 佘健俊; 李梅

    2013-01-01

    精益建造是一种先进的生产管理模式,对新时期建筑业企业精益建造能力的评价研究有助于建筑业企业改进施工生产模式,真正做到精益采购、精益施工以及精益交付。根据精益价值链的特点和要求建立了一套建筑业企业精益建造能力评价指标体系,其中重点对采购流程中的准时采购、按需采购和施工流程中的质量管理以及消除浪费等指标进行了分析,利用模糊层次分析法对建筑业企业的精益建造能力进行综合评价,并进行实证分析,以期为建筑业企业精益建造能力的评价与提升提供一种借鉴方法。%Lean construction is an advanced production management mode. The research of the evaluation of construction enterprise’s lean construction ability in the new period can help construction enterprises to improve production mode,and to realize lean procurement,lean construction and lean delivery. The paper firstly introduces the concept and characteristics of lean value chain,and then creates a set of index system of construction enterprise’s lean construction ability according to the requirements of the lean value chain. Specifically,the paper focuses on analyzing some index,such as just in time purchasing,on-demand procurement in procurement process,as well as quality management and waste elimination in the construction process,then conducting a comprehensive evaluation of construction enterprise’s lean ability by fuzzy analytic hierarchy process,and finally the paper carries on an empirical analysis,which provides a reference method for the evaluation and improvement of construction enterprise’s lean construction ability.

  20. ImmunoPET imaging of B-cell lymphoma using 124I-anti-CD20 scFv dimers (diabodies).

    Science.gov (United States)

    Olafsen, Tove; Sirk, Shannon J; Betting, David J; Kenanova, Vania E; Bauer, Karl B; Ladno, Waldemar; Raubitschek, Andrew A; Timmerman, John M; Wu, Anna M

    2010-04-01

    Rapid clearing engineered antibody fragments for immunoPET promise high sensitivity at early time points. Here, tumor targeting of anti-CD20 diabodies (scFv dimers) for detection of low-grade B-cell lymphomas were evaluated. In addition, the effect of linker length on oligomerization of the diabody was investigated. Four rituximab scFv variants in the V(L)-V(H) orientation with different linker lengths between the V domains (scFv-1, scFv-3, scFv-5, scFv-8), plus the scFv-5 with a C-terminal cysteine (Cys-Db) for site-specific modification were generated. The scFv-8 and Cys-Db were radioiodinated with (124)I for PET imaging, and biodistribution of (131)I-Cys-Db was carried out at 2, 4 10 and 20 h. The five anti-CD20 scFv variants were expressed as fully functional dimers. Shortening the linker to three or one residue did not produce higher order of multimers. Both (124)I-labeled scFv-8 and Cys-Db exhibited similar tumor targeting at 8 h post injection, with significantly higher uptakes than in control tumors (P < 0.05). At 20 h, less than 1% ID/g of (131)I-labeled Cys-Db was present in tumors and tissues. Specific tumor targeting and high contrast images were achieved with the anti-CD20 diabodies. These agents extend the repertoire of reagents that can potentially be used to improve detection of low-grade lymphomas. PMID:20053640

  1. High-level production in Pichia pastoris of an anti-p185HER-2 single-chain antibody fragment using an alternative secretion expression vector.

    Science.gov (United States)

    Gurkan, Cemal; Symeonides, Stefan N; Ellar, David J

    2004-02-01

    The methylotrophic yeast Pichia pastoris has become a highly popular expression host for the recombinant production of a wide variety of proteins. Initial success with this system was greatly facilitated by the development of versatile expression vectors that were almost exclusively based on the strong, tightly regulated promoter of the P. pastoris major alcohol oxidase gene ( AOX1 ). For example, pIB4 is an Escherichia coli - P. pastoris shuttle vector that also uses the AOX1 promoter to allow intracellular expression of endogenous and foreign genes in the latter organism. Since the eukaryotic advantages of P. pastoris would be best harnessed through the secretory targeting of the recombinant proteins, we modified the pIB4 vector by adding the Saccharomyces cerevisiae alpha-factor secretion signal immediately upstream of its multiple cloning site. Here we describe the construction of this modified vector, pIB4alpha, and its successful use for the high-level expression and secretion of a functional single-chain antibody fragment (scFv), C6.5, which targets p185(HER-2), a cell-surface glycoprotein overexpressed in about 30% of human breast and ovarian cancers. The PCR strategy used for the subcloning of the C6.5 construct into pIB4alpha also introduced a short DNA sequence coding for a C-terminal hexahistidine tag, which allowed subsequent purification of the secreted scFv, by immobilized-metal-affinity chromatography, to a yield of 70 mg x l(-1) of shake-flask culture. In conclusion, our results suggest that the secretion expression vector pIB4alpha not only complements the original pIB4 vector for intracellular expression in P. pastoris, but might also constitute an attractive alternative to the commercially available secretion expression vectors. PMID:12962542

  2. Research on Enterprise Social Responsibility Construction in Supply Chain Based on the Perspectives of Network Governance%基于网络治理视角的供应链企业社会责任研究

    Institute of Scientific and Technical Information of China (English)

    周翼翔

    2015-01-01

    At present, corporate social responsibility is no longer confined to the scope of an enterprise , but extended to different links of supply chain.In supply chain, the position, profit distribution, and corporate social responsibility performance of each enterprise are not identical, which makes both core and non-core enterprises lack of responsibility.According to the situation of"prisoner's dilemma", we construct the model to pro-mote corporate social responsibility in supply chain .Only from point to chain and then to network governance can we gradually change the current situation of deficient corporate social responsibility on the basis of stimulus , contract constraint, mutual competition combined with external govern-ance in our country.%企业社会责任的范围已不再局限于某个企业,已扩展到供应链的不同环节。在供应链中,由于各企业的地位、利益分配以及企业社会责任表现各不相同,使得无论核心企业还是非核心企业都缺乏履行责任的动力。针对这种“囚徒困境”的局面,构建供应链企业社会责任推进模式,在经济刺激、协议约束、相互竞争的基础上结合外部治理,由点到链,再由链到网络治理逐步推进来解决我国目前企业社会责任缺失的现状。

  3. A chain of FPU cells

    OpenAIRE

    Verhulst, F.

    2015-01-01

    In contrast to the classical Fermi-Pasta-Ulam (FPU) chain, the inhomogeneous FPU chain shows nearly all the principal resonances. Using this fact, we can construct a periodic FPU chain of low dimension, for instance a FPU cell of four degrees-of-freedom, that can be used as a building block for a chain of FPU cells. Differences between chains in nearest-neighbour interaction and those in overall interaction are caused by symmetry. We will also show some results on the dynamics of a particular...

  4. Dynamics of corporate strategy from a value chain perspective : A study of the Swedish telecom and construction industries during the 90’s

    OpenAIRE

    de Paula, Andes

    2006-01-01

    Changes in sectors and industries have brought new challenges to corporations as well as been important driving forces for the dynamics in strategy at the corporate level. With the dramatic developments of the 1990’s in mind, such as multilateral free-trade agreements, liberalization, privatization, sharp industry growth/decline, increased competition and globalization, in particular within the telecom and the construction industry, this study contributes to describing and understanding strat...

  5. Insight into earthquake sequencing: analysis and interpretation of time-series constructed from the directed graph of the Markov chain model

    Directory of Open Access Journals (Sweden)

    M. S. Cavers

    2015-02-01

    Full Text Available Directed graph representation of a Markov chain model to study global earthquake sequencing leads to a time-series of state-to-state transition probabilities that includes the spatio-temporally linked recurrent events in the record-breaking sense. A state refers to a configuration comprised of zones with either the occurrence or non-occurrence of an earthquake in each zone in a pre-determined time interval. Since the time-series is derived from non-linear and non-stationary earthquake sequencing, we use known analysis methods to glean new information. We apply decomposition procedures such as ensemble empirical mode decomposition (EEMD to study the state-to-state fluctuations in each of the intrinsic mode functions. We subject the intrinsic mode functions, the orthogonal basis set derived from the time-series using the EEMD, to a detailed analysis to draw information-content of the time-series. Also, we investigate the influence of random-noise on the data-driven state-to-state transition probabilities. We consider a second aspect of earthquake sequencing that is closely tied to its time-correlative behavior. Here, we extend the Fano factor and Allan factor analysis to the time-series of state-to state transition frequencies of a Markov chain. Our results support not only the usefulness the intrinsic mode functions in understanding the time-series but also the presence of power-law behaviour exemplified by the Fano factor and the Allan factor.

  6. Characterization of single chain antibody targets through yeast two hybrid

    Directory of Open Access Journals (Sweden)

    Vielemeyer Ole

    2010-08-01

    Full Text Available Abstract Background Due to their unique ability to bind their targets with high fidelity, antibodies are used widely not only in biomedical research, but also in many clinical applications. Recombinant antibodies, including single chain variable fragments (scFv, are gaining momentum because they allow powerful in vitro selection and manipulation without loss of function. Regardless of the ultimate application or type of antibody used, precise understanding of the interaction between the antibody's binding site and its specific target epitope(s is of great importance. However, such data is frequently difficult to obtain. Results We describe an approach that allows detailed characterization of a given antibody's target(s using the yeast two-hybrid system. Several recombinant scFv were used as bait and screened against highly complex cDNA libraries. Systematic sequencing of all retained clones and statistical analysis allowed efficient ranking of the prey fragments. Multiple alignment of the obtained cDNA fragments provided a selected interacting domain (SID, efficiently narrowing the epitope-containing region. Interactions between antibodies and their respective targets were characterized for several scFv. For AA2 and ROF7, two conformation-specific sensors that exclusively bind the activated forms of the small GTPases Rab6 and Rab1 respectively, only fragments expressing the entire target protein's core region were retained. This strongly suggested interaction with a non-linear epitope. For two other scFv, TA10 and SF9, which recognize the large proteins giantin and non-muscle myosin IIA, respectively, precise antibody-binding regions within the target were defined. Finally, for some antibodies, secondary targets within and across species could be revealed. Conclusions Our method, utilizing the yeast two-hybrid technology and scFv as bait, is a simple yet powerful approach for the detailed characterization of antibody targets. It allows precise

  7. Construction of a human antibody domain (VH) library

    OpenAIRE

    Chen, Weizao; Zhu, Zhongyu; Xiao, Xiaodong; Dimitrov, Dimiter S.

    2009-01-01

    Highly diverse antibody (Fab or scFv) libraries have become vital sources to select antibodies with high affinity and novel properties. Combinatorial strategies provide efficient ways of creating antibody libraries containing a large number of individual clones. These strategies include the reassembly of naturally occurring genes encoding the heavy and light chains from either immune or nonimmune B-cell sources, or introduction of synthetic diversity to either the framework regions (FRs) or t...

  8. Model protocells from single-chain lipids.

    Science.gov (United States)

    Mansy, Sheref S

    2009-03-01

    Significant progress has been made in the construction of laboratory models of protocells. Most frequently the developed vesicle systems utilize single-chain lipids rather than the double-chain lipids typically found in biological membranes. Although single-chain lipids yield less robust vesicles, their dynamic characteristics are highly exploitable for protocellular functions. Herein the advantages of using single-chain lipids in the construction of protocells are discussed. PMID:19399223

  9. Model Protocells from Single-Chain Lipids

    OpenAIRE

    Mansy, Sheref S.

    2009-01-01

    Significant progress has been made in the construction of laboratory models of protocells. Most frequently the developed vesicle systems utilize single-chain lipids rather than the double-chain lipids typically found in biological membranes. Although single-chain lipids yield less robust vesicles, their dynamic characteristics are highly exploitable for protocellular functions. Herein the advantages of using single-chain lipids in the construction of protocells are discussed.

  10. Model Protocells from Single-Chain Lipids

    Directory of Open Access Journals (Sweden)

    Sheref S. Mansy

    2009-03-01

    Full Text Available Significant progress has been made in the construction of laboratory models of protocells. Most frequently the developed vesicle systems utilize single-chain lipids rather than the double-chain lipids typically found in biological membranes. Although single-chain lipids yield less robust vesicles, their dynamic characteristics are highly exploitable for protocellular functions. Herein the advantages of using single-chain lipids in the construction of protocells are discussed.

  11. Recovery of active anti TNF-α ScFv through matrix-assisted refolding of bacterial inclusion bodies using CIM monolithic support.

    Science.gov (United States)

    Sushma, Krishnan; Bilgimol, Chuvappumkal Joseph; Vijayalakshmi, Mookambeswaran A; Satheeshkumar, Padikara Kutty

    2012-04-01

    Anti TNF-α molecules are important as therapeutic agents for many of the autoimmune diseases in chronic stage. Here we report the expression and purification of a recombinant single chain variable fragment (ScFv) specific to TNF-α from inclusion bodies. In contrast to the conventional on column refolding using the soft gel supports, an efficient methodology using monolithic matrix has been employed. Nickel (II) coupled to convective interaction media (CIM) support was utilized for this purpose with 6M guanidine hydrochloride (GuHCl) as the chaotropic agent. The protein purified after solubilization and refolding proved to be biologically active with an IC₅₀ value of 15 μg. To the best of our knowledge, this is the first report showing the application of methacrylate based chromatographic supports for matrix-assisted refolding and purification of Escherichia coli inclusion bodies. The results are promising to elaborate the methodology further to exploit the potential positive features of monoliths in protein refolding science. PMID:22386363

  12. FV-429 induces apoptosis and inhibits glycolysis by inhibiting Akt-mediated phosphorylation of hexokinase II in MDA-MB-231 cells.

    Science.gov (United States)

    Zhou, Yuxin; Lu, Na; Qiao, Chen; Ni, Ting; Li, Zhiyu; Yu, Boyang; Guo, Qinglong; Wei, Libin

    2016-09-01

    In this study, the anticancer effect of a newly synthesized flavonoid FV-429, against human breast cancer MDA-MB-231 cells, and the underlying mechanisms were investigated. FV-429 triggered the apoptosis and simultaneously inhibited the glycolysis of MDA-MB-231 cells. Both the HK II activity and its level in mitochondria were significantly down regulated by FV-429. Moreover, FV-429 weakened the interaction between HKII and VDAC, stimulated the detachment of HK II from the mitochondria, and resulted in the opening of the mitochondrial permeability transition pores. Thus FV-429 induced the mitochondrial-mediated apoptosis, showing increased Bax/Bcl-2 ratio, loss of mitochondrial membrane potential (MMP) and activation of caspase-3 and -9, cytochrome c (Cyt c) release, and apoptosis inducing factor (AIF) transposition. Further research revealed that the phosphorylation of mitochondrial HKII via Akt was responsible for the dissociation of HKII and the decreased HKII activity induced by FV-429. Taken together, FV-429 inhibited the phosphorylation of HKII, down-regulated its activity, and stimulated the release of HKII from the mitochondria, resulting the inhibited glycolysis and mitochondrial-mediated apoptosis. The studies provide a molecular basis for the development of flavonoid compounds as novel anticancer agents for breast cancer. © 2015 Wiley Periodicals, Inc. PMID:26258875

  13. Anti-Staphylococcus aureus single-chain variable region fragments provide protection against mastitis in mice.

    Science.gov (United States)

    Wang, Man; Zhang, Yan; Zhu, Jianguo

    2016-03-01

    Staphylococcus aureus is a leading causative agent of bovine mastitis, which can result in significant economic losses to the dairy industry. However, available vaccines against bovine mastitis do not confer adequate protection, although passive immunization with antibodies may be useful to prevent disease. Hence, we constructed a bovine single-chain variable region fragment (scFv) phage display library using cDNAs from peripheral blood lymphocytes of cows with S. aureus-induced mastitis. After four rounds of selection, eight scFvs that bound S. aureus antigens with high affinity were obtained. The framework regions of the variable domains (VH and VL) of the eight scFvs were highly conserved, and the complementarity-determining regions (CDRs) displayed significant diversity, especially CDR3 of the VH domain. All eight scFvs inhibited S. aureus growth in culture medium. Lactating mice were challenged by injecting S. aureus into the fourth mammary gland. Histopathological analysis showed that treatment with these scFvs prior to bacterial challenge maintained the structure of the mammary acini, decreased infiltration of polymorphonuclear neutrophils, increased levels of interferon-gamma and interleukin-4, and reduced tumor necrosis factor-alpha levels in mammary tissues, as compared with mice treatment with physiological saline (P < 0.05). These novel bovine scFvs may be suitable candidates for therapeutic agents for the prevention of S. aureus-induced bovine mastitis. PMID:26512007

  14. Research on Integrated Green Supply Chain Management

    Institute of Scientific and Technical Information of China (English)

    DENG Lei; WANG Xu

    2006-01-01

    On the basis of the analyzing product life cycle and value chain management in green supply chain, integrated green supply chain is put forward and constructed which involve lean production, agile manufacturing and green supply chain. This integrated structure provides an effective method for resolving some questions such as cost, market, environment, etc. in enterprise. A case study is presented at the end of paper to demonstrate how integrated supply chain implemented successfully in enterprise.

  15. Wheat cultivars selected for high Fv/Fm under heat stress maintain high photosynthesis, total chlorophyll, stomatal conductance, transpiration and dry matter

    DEFF Research Database (Denmark)

    Sharma, Dew Kumari; Andersen, Sven Bode; Ottosen, Carl-Otto;

    2015-01-01

    The chlorophyll fluorescence parameter Fv/Fm reflects the maximum quantum efficiency of photosystem II (PSII) photochemistry and has been widely used for early stress detection in plants. Previously, we have used a three-tiered approach of phenotyping by Fv/Fm to identify naturally existing genetic......) than the low group, accompanied by higher stomatal conductance (gs), transpiration rate (E) and evaporative cooling of the leaf (ΔT). The difference in PN between the groups was not caused by differences in PSII capacity or gs as the variation in Fv/Fm and intracellular CO2 (Ci) was non...

  16. Generation of recombinant single-chain antibodies neutralizing the cytolytic activity of vaginolysin, the main virulence factor of Gardnerella vaginalis

    Directory of Open Access Journals (Sweden)

    Pleckaityte Milda

    2011-11-01

    Full Text Available Abstract Background Gardnerella vaginalis is identified as the predominant colonist of the vaginal tract in women with bacterial vaginosis. Vaginolysin (VLY is a protein toxin released by G. vaginalis. VLY possesses cytolytic activity and is considered as a main virulence factor of G. vaginalis. Inhibition of VLY-mediated cell lysis by antibodies may have important physiological relevance. Results Single-chain variable fragments of immunoglobulins (scFvs were cloned from two hybridoma cell lines producing neutralizing antibodies against VLY and expressed as active proteins in E. coli. For each hybridoma, two variants of anti-VLY scFv consisting of either VL-VH or VH-VL linked with a 20 aa-long linker sequence (G4S4 were constructed. Recovery of scFvs from inclusion bodies with subsequent purification by metal-chelate chromatography resulted in VLY-binding proteins that were predominantly monomeric. The antigen-binding activity of purified scFvs was verified by an indirect ELISA. The neutralizing activity was investigated by in vitro hemolytic assay and cytolytic assay using HeLa cell line. Calculated apparent Kd values and neutralizing potency of scFvs were in agreement with those of parental full-length antibodies. VH-VL and VL-VH variants of scFvs showed similar affinity and neutralizing potency. The anti-VLY scFvs derived from hybridoma clone 9B4 exhibited high VLY-neutralizing activity both on human erythrocytes and cervical epithelial HeLa cells. Conclusions Hybridoma-derived scFvs with VLY-binding activity were expressed in E. coli. Recombinant anti-VLY scFvs inhibited VLY-mediated cell lysis. The monovalent scFvs showed reduced affinity and neutralizing potency as compared to the respective full-length antibodies. The loss of avidity could be restored by generating scFv constructs with multivalent binding properties. Generated scFvs is the first example of recombinant single-chain antibodies with VLY-neutralizing activity produced in

  17. 基于建筑供应链的不允许缺货联合库存模型研究%Study on Joint Managed Inventory Model Under No-Shortage Based on Construction Materials Supply Chain

    Institute of Scientific and Technical Information of China (English)

    钟德强; 郭薇

    2014-01-01

    研究由一个总承包商与多个分包商组成的建筑材料供应链联合库存问题,在不允许缺货的情况下,建立了库存成本模型。传统独立库存与供应链联合库存的成本比较证明,供应链联合库存总成本低于传统独立库存总成本。并以一个总承包商、2个分包商为例,研究了联合库存下的成本分配问题,证明用Shapley值法分配成本,各参与方在供应链联合库存下分担的成本均低于传统独立库存下的成本。%Investigates the joint managed inventory(JMI) problems of construction materials supply chain consist-ing of one general contractor and multiple sub-contractors. On the condition that short supply is not allowed,establishes the inventory cost model. Comparing the traditional independent inventory cost and that of supply chain JMI, proves that the total cost of the JMI is lower. Taking one general contractor and two sub-contractors for an example, studies the cost allocation of joint managed inventory, and verifies that using Shapley value method to allocate the cost, the shared cost of each participant in supply chain JMI is lower than that of traditional independent inventory.

  18. Synthesis, crystal structure and properties of two 1D nano-chain coordination polymers constructed by lanthanide with pyridine-3,4-dicarboxylic acid and 1,10-phenanthroline

    International Nuclear Information System (INIS)

    The hydrothermal reactions of LnCl3.6H2O (Ln=Eu, Tb), pyridine-3,4-dicarboxylic acid (3,4-pydaH2), 1,10-phenthroline (phen) and NaOH in aqueous medium yield two metal-organic hybrid materials, [Eu2(3,4-pyda)3(phen)(H2O).H2O]n (1) and [Tb2(3,4-pyda)3(phen)(H2O).H2O]n (2), respectively. Both compounds have similar topology structure containing one-dimensional nano-chain, which is further assembled into a three-dimensional supramolecular network via π-π stacking interactions and hydrogen bonds. To the best of our knowledge, they represent the first example of nano-chain coordination polymers constructed by 3,4-pydaH2 and chelate heterocylic ligand. Interestingly, the 3,4-pyda anion exhibits three kinds of coordination modes in these complexes. The coordination modes of 3,4-pyda in complexes 1 and 2 have not been observed in other coordination polymers containing 3,4-pyda ligands. Compounds 1 and 2 exhibit strong fluorescent emission bands in the solid state at room temperature. Their magnetic analyses show that they exhibit different magnetic interactions. - Graphical abstract: Two novel lanthanide coordination polymers [M2(pydc)3(phen)(H2O).H2O]n (M=Eu(1) and Tb(2), pydc=pyridine-3,4-dicarboxylate, phen=1,10-phenthroline) have been synthesized and characterized. Both compounds reveal a one-dimensional nano-chain, which is further assembled into a three-dimensional supramolecular network via π-π stacking interactions and hydrogen bonds. Their luminescent and magnetic properties have been investigated

  19. 物联网环境下建筑供应链库存协同管理的研究%Study on Construction Supply Chain Collaborative Inventory Management in IOT Environment

    Institute of Scientific and Technical Information of China (English)

    王连月

    2014-01-01

    In this paper, through illustrating the connotation of the collaborative inventory management of the construction supply chain, we analyzed the problems therein and pointed out that in order to realize the collaborative inventory management for the supply chain, the IOT network and inventory management information platform of the nodal enterprises should be set up first.%为了实现建筑供应链库存协同管理,需要各节点企业数据准确传输和信息充分共享。通过对建筑供应链库存协同管理内涵的论述,分析了供应链库存协同管理的问题,提出实现供应链库存协同管理需要构建节点企业物联网和库存管理信息平台。

  20. The production of recombinant single chain antibody fragments for the detection of illicit drug residues

    OpenAIRE

    Brennan, Joanne

    2005-01-01

    Recombinant antibodies represent a more sensitive and specific detection tool for immunoanalysis. The research carried out for this thesis describes the production of genetically-derived single chain antibody fragments to detect illicit drugs. A variety of novel recombinant antibody fragments against morphine-3-glucuronide, a metabolite of heroin has been produced. A monomeric, dimeric and enzyme-labelled scFv were characterised with respect to their binding abilities and cross reactiviti...

  1. Single-Chain Fragment Variable Passive Immunotherapies for Neurodegenerative Diseases

    OpenAIRE

    Liang Huang; Federoff, Howard J.; Xiaomin Su

    2013-01-01

    Accumulation of misfolded proteins has been implicated in a variety of neurodegenerative diseases including prion diseases, Alzheimer’s disease (AD), Parkinson’s disease (PD), and Huntington’s disease (HD). In the past decade, single-chain fragment variable (scFv) -based immunotherapies have been developed to target abnormal proteins or various forms of protein aggregates including Aβ, SNCA, Htt, and PrP proteins. The scFvs are produced by fusing the variable regions of the antibody heavy and...

  2. Construction and characterization of a non-immune Llama variable heavy chain phage display antibody library%羊驼非免疫重链单域抗体库的构建和鉴定

    Institute of Scientific and Technical Information of China (English)

    吴标; 王树军; 夏立亮; 季萍; 葛海良; 赵国屏; 王颖

    2011-01-01

    本研究旨在通过构建羊驼非免疫重链单域抗体库,完成抗体库多样性的鉴定,为进一步筛选抗原特异性重链抗体奠定基础.我们从未经免疫的羊驼外周血中分离外周血单个核细胞(PBMC),抽提RNA后,用RT-PCR方法特异性扩增羊驼重链抗体可变区(VHH)片段;并采用两步连接方法将重链抗体可变区片段与噬菌粒载体pCANTAB5E连接获得重组子,多次电转感受态大肠杆菌TG1后获得VHH抗体基因库;并采用稀释计数法测定抗体库库容量,随机挑取克隆测序验证抗体库多样性.结果显示,我们所构建的羊驼非免疫重链单域抗体库的库容量为1.5×109,随机克隆测序验证多样性良好,独立克隆所占比例为80%,并显示出和人源抗体较高的同源性.上述结果表明,我们已经成功构建获得大容量的羊驼非免疫重链单域抗体库,为进一步筛选抗原特异性重链抗体奠定基础.%To construct a non-immune Llama variable domain of heavy chain antibody(VHH) phage display antibody library (VHH antibody library). Llama peripheral blood mononuclear lymphocytes were isolated from whole blood by Ficoll-hypaque density gradient centrifugation. Total RNA was extracted from PBMCs and reverse-transcripted into cDNA by using specific primers. VHH were amplified by nested PCR. PCR products of VHH fragments were then purified and ligated with phagemid vector pCANTAB5E by a modified two-step ligation method. Recombinant pCANTAB5E-VHH vectors were electroporated into competent TGI E.coli cells to obtain the primary VHH antibody library. The library capacity was titrated through limited dilution. Recombination efficacy and diversity of VHH antibody library was determined by sequencing analysis. Alignment a-nalysis was performed to compare the homology between Llama VHH domain and human/mouse variable regions of heavy chains. By using a modified strategy, we have constructed a non-immune Llama VHH antibody library with 1. 5

  3. 基于供应链的应急物流虚拟联合体的构建%The Construction of Emergency Logistics Virtual Union Based on Supply Chain

    Institute of Scientific and Technical Information of China (English)

    陈伟珂; 花翠

    2014-01-01

    This paper based on supply chain management theory,follows the point -line -surface construction principle, proposed to construct a framework of emergency logistics virtual consortium,in order to give full play to the leading role of the government,the army vanguard role,the market supply,the practical realization of military logistics and logistics,the organic combination of market mechanism and administration mechanism.The purpose is to effectively integrate social and professional resources,improve the overall response speed and supply efficiency of emergency logistics.%文中基于供应链管理理论,遵循点-线-面构建原则,提出应急物流虚拟联合体的构建框架,以充分发挥政府主导作用,军队先锋作用,市场供给作用,切实实现军队物流和地方物流、市场机制和行政机制的有机结合。目的是有效集成社会专业资源,提高应急物流的整体响应速度和供应效率。

  4. Improving corrosion and wear resistance of FV520B steel by high current pulsed electron beam surface treatment

    International Nuclear Information System (INIS)

    Highlights: • HCPEB surface treatment was conducted on FV520B steel. • Surface layer ∼10 μm was refined with elements homogenization. • Remelted layer exhibited 〈2 0 0〉 preferential orientation. • Corrosion potential increased and corrosion rate decreased one order of magnitude. • Wear resistance increased by 3 times. - Abstract: High current pulsed electron beam (HCPEB) surface treatment was conducted on FV520B steel with accelerating voltage 27 kV, pulse duration 2.5 μs, energy density 5 J/cm2 and 1–25 pulses. The surface microstructure and element distribution were examined by using scanning electron microscope (SEM), electron probe micro-analysis (EPMA) and X-ray diffraction (XRD) methods. After HCPEB treatments, the surface microstructure became refined and uniform with an average grain size less than 2 μm and a preferential solidification orientation in 〈2 0 0〉 direction. The thickness of surface remelted layer was ∼4 μm. The initial precipitated particles in surface layer of depth ∼10 μm were dissolved into the base matrix and gave a homogenous element distribution. The HCPEB modified surface exhibited an effective improvement in corrosion and wear resistance. The corrosion potential shifted towards positive and the corrosion rate decreased nearly one order of magnitude, while the wear resistance after 25 pulses of HCPEB treatment increased by 3 times as compared with the initial FV520B steel

  5. Synthesis and pre-clinical evaluation of an (18)F-labeled single-chain antibody fragment for PET imaging of epithelial ovarian cancer.

    Science.gov (United States)

    Sharma, Sai Kiran; Wuest, Melinda; Way, Jenilee D; Bouvet, Vincent R; Wang, Monica; Wuest, Frank R

    2016-01-01

    Anti-CA125 antibodies have been used in immunoassays to quantify levels of shed antigen in the serum of patients who are under surveillance for epithelial ovarian cancer (EOC). However, there is currently no molecular imaging probe in the clinic for the assessment of CA125 expression in vivo. The present study describes the development of an (18)F-labeled single-chain variable fragment (scFv) for PET imaging of CA125 in preclinical EOC models. Anti-CA125 scFv was derived from MAb-B43.13 by recombinant expression of the fragment in E.coli. Fragment scFv-B43.13 was purified via immobilized metal affinity chromatography and characterized for antigen binding via immuno-staining and flow cytometry. Prosthetic group N-succinimidyl 4-[(18)F]fluorobenzoate ([(18)F]SFB) was used for radiolabeling of scFv-B43.13. Preclinical ovarian cancer models were developed based on ovarian cancer cell lines OVCAR3 (CA125-positive) and SKOV3 (CA125-negative) in NIH-III mice. The radiopharmacological profile of (18)F-labeled scFv-B43.13 ([(18)F]FBz-scFv-B43.13) was studied with PET. [(18)F]FBz-scFv-B43.13 was prepared in radiochemical yields of 3.7 ± 1.8% (n = 5) at an effective specific activity of 3.88 ± 0.76 GBq/µmol (n = 5). The radiotracer demonstrated selective uptake in CA125-positive OVCAR3 cells and virtually no uptake in CA125-negative SKOV3 cells. Standardized uptake values (SUV) of radioactivity uptake in OVCAR3 tumors was 0.5 (n = 3) and 0.3 (n = 2) in SKOV3 tumors after 60 min post injection (p.i.). PMID:27508105

  6. Synthesis and pre-clinical evaluation of an 18F-labeled single-chain antibody fragment for PET imaging of epithelial ovarian cancer

    Science.gov (United States)

    Sharma, Sai Kiran; Wuest, Melinda; Way, Jenilee D; Bouvet, Vincent R; Wang, Monica; Wuest, Frank R

    2016-01-01

    Anti-CA125 antibodies have been used in immunoassays to quantify levels of shed antigen in the serum of patients who are under surveillance for epithelial ovarian cancer (EOC). However, there is currently no molecular imaging probe in the clinic for the assessment of CA125 expression in vivo. The present study describes the development of an 18F-labeled single-chain variable fragment (scFv) for PET imaging of CA125 in preclinical EOC models. Anti-CA125 scFv was derived from MAb-B43.13 by recombinant expression of the fragment in E.coli. Fragment scFv-B43.13 was purified via immobilized metal affinity chromatography and characterized for antigen binding via immuno-staining and flow cytometry. Prosthetic group N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB) was used for radiolabeling of scFv-B43.13. Preclinical ovarian cancer models were developed based on ovarian cancer cell lines OVCAR3 (CA125-positive) and SKOV3 (CA125-negative) in NIH-III mice. The radiopharmacological profile of 18F-labeled scFv-B43.13 ([18F]FBz-scFv-B43.13) was studied with PET. [18F]FBz-scFv-B43.13 was prepared in radiochemical yields of 3.7 ± 1.8% (n = 5) at an effective specific activity of 3.88 ± 0.76 GBq/µmol (n = 5). The radiotracer demonstrated selective uptake in CA125-positive OVCAR3 cells and virtually no uptake in CA125-negative SKOV3 cells. Standardized uptake values (SUV) of radioactivity uptake in OVCAR3 tumors was 0.5 (n = 3) and 0.3 (n = 2) in SKOV3 tumors after 60 min post injection (p.i.). PMID:27508105

  7. The radiosensitizing properties of an anti-epidermal growth factor receptor single-chain antibody isolated from a phage display library

    International Nuclear Information System (INIS)

    Full text: Anti-epidermal growth factor receptor (EGFr) agents have shown promise in the treatment of various malignancies when used as single agents or combined with conventional treatments. These agents include monoclonal antibodies that block the ligand binding site of EGFr. The studies reported herein were performed to isolate single-chain antibodies (scFvs) that target EGFr and to characterize the anti-cancer efficacy of these smaller antibody molecules. It is our hypothesis that therapeutically effective anti-EGFr scFvs could eventually be delivered in a gene-therapy approach that allows affected tumor cells to secrete the anti-EGFr scFvs thereby impacting multiple neighboring cells. Human scFv phage display libraries were screened for EGFr-binding scFvs. One positive EGFr-specific scFv (clone 45) was tested for its ability to sensitize tumor cells to radiation treatment. The EGFr-over expressing cell line, A431 cells (human squamous cell carcinoma), was used in standard cell proliferation and apoptosis (annexin V) assays. A431 cells were treated with EGFr-specific scFv clone 45 (50 μg/ml), 3 Gy or the combination of the two treatments. Cell proliferation was assessed daily and all treatments inhibited proliferation, however; greater inhibition of cell proliferation was noted for the combination treatment than either individual treatment. Inhibition at 4 days compared to controls: 26% (scFv), 32% (3 Gy), and 54% (combined). Cells treated in a similar fashion were studied for apoptosis 4 days after the initiation of treatment. Although the scFv did not induce apoptosis, it did cause a significant increase in radiation-induced apoptosis. An scFv was isolated from a human scFv phage display library and shown to sensitize human A431 cells to radiation treatment. Further studies to determine the mechanism of radiosensitization are being undertaken

  8. Anti-CD20 single chain variable antibody fragment–apolipoprotein A-I chimera containing nanodisks promote targeted bioactive agent delivery to CD20-positive lymphomas

    Science.gov (United States)

    Crosby, Natasha M.; Ghosh, Mistuni; Su, Betty; Beckstead, Jennifer A.; Kamei, Ayako; Simonsen, Jens B.; Luo, Bing; Gordon, Leo I.; Forte, Trudy M.; Ryan, Robert O.

    2015-01-01

    A fusion protein comprising an α-CD20 single chain variable fragment (scFv) antibody, a spacer peptide, and human apolipoprotein (apo) A-I was constructed and expressed in Escherichia coli. The lipid interaction properties intrinsic to apoA-I as well as the antigen recognition properties of the scFv were retained by the chimera. scFv•apoA-I was formulated into nanoscale reconstituted high-density lipoprotein particles (termed nanodisks; ND) and incubated with cultured cells. α-CD20 scFv•apoA-I ND bound to CD20-positive non-Hodgkins lymphoma (NHL) cells (Ramos and Granta) but not to CD20-negative T lymphocytes (i.e., Jurkat). Binding to NHL cells was partially inhibited by pre-incubation with rituximab, a monoclonal antibody directed against CD20. Confocal fluorescence microscopy analysis of Granta cells following incubation with α-CD20 scFv•apoA-I ND formulated with the intrinsically fluorescent hydrophobic polyphenol, curcumin, revealed α-CD20 scFv•apoA-I localizes to the cell surface, while curcumin off-loads and gains entry to the cell. Compared to control incubations, viability of cultured NHL cells was decreased upon incubation with α-CD20 scFv•apoA-I ND harboring curcumin. Thus, formulation of curcumin ND with α-CD20 scFv•apoA-I as the scaffold component confers cell targeting and enhanced bioactive agent delivery, providing a strategy to minimize toxicity associated with chemotherapeutic agents. PMID:25994015

  9. Anti-CD20 single chain variable antibody fragment-apolipoprotein A-I chimera containing nanodisks promote targeted bioactive agent delivery to CD20-positive lymphomas.

    Science.gov (United States)

    Crosby, Natasha M; Ghosh, Mistuni; Su, Betty; Beckstead, Jennifer A; Kamei, Ayako; Simonsen, Jens B; Luo, Bing; Gordon, Leo I; Forte, Trudy M; Ryan, Robert O

    2015-08-01

    A fusion protein comprising an α-CD20 single chain variable fragment (scFv) antibody, a spacer peptide, and human apolipoprotein (apo) A-I was constructed and expressed in Escherichia coli. The lipid interaction properties intrinsic to apoA-I as well as the antigen recognition properties of the scFv were retained by the chimera. scFv•apoA-I was formulated into nanoscale reconstituted high-density lipoprotein particles (termed nanodisks; ND) and incubated with cultured cells. α-CD20 scFv•apoA-I ND bound to CD20-positive non-Hodgkins lymphoma (NHL) cells (Ramos and Granta) but not to CD20-negative T lymphocytes (i.e., Jurkat). Binding to NHL cells was partially inhibited by pre-incubation with rituximab, a monoclonal antibody directed against CD20. Confocal fluorescence microscopy analysis of Granta cells following incubation with α-CD20 scFv•apoA-I ND formulated with the intrinsically fluorescent hydrophobic polyphenol, curcumin, revealed α-CD20 scFv•apoA-I localizes to the cell surface, while curcumin off-loads and gains entry to the cell. Compared to control incubations, viability of cultured NHL cells was decreased upon incubation with α-CD20 scFv•apoA-I ND harboring curcumin. Thus, formulation of curcumin ND with α-CD20 scFv•apoA-I as the scaffold component confers cell targeting and enhanced bioactive agent delivery, providing a strategy to minimize toxicity associated with chemotherapeutic agents. PMID:25994015

  10. Antitumor effects of an engineered and energized fusion protein consisting of an anti-CD20 scFv fragment and lidamycin.

    Science.gov (United States)

    Fang, Hong; Miao, Qingfang; Zhang, Shenghua; Cheng, Xin; Xiong, Dongsheng; Zhen, Yongsu

    2011-03-01

    Antibody-based fusion proteins are the next generation of antibody therapies for cancer and other diseases. CD20 antigen, which is overexpressed on cell membranes in nearly 95% of cases of B-cell Non-Hodgkin's Lymphoma, is an attractive target for the therapy of B-lymphoid malignancies. Lidamycin (LDM) is a potent enediyne-containing antitumor antibiotic that now has entered phase II clinical trials. In this study, we prepared an engineered fusion protein, scFv-LDP, consisting of an anti-CD20 scFv fragment and the apoprotein LDP of LDM using DNA recombination. After purification and refolding, scFv-LDP was found to bind specifically to CD20-positive lymphoma cells using ELISA and indirect immunofluorescent cytochemical staining assays. The energized fusion protein scFv-LDP-AE was obtained using molecular reconstitution of the active chromophore AE of LDM and scFv-LDP. MTT assay revealed potent cytotoxicity of scFv-LDP-AE to CD20-positive Raji and Daudi cells, with IC(50) values of 1.21×10(-11) and 6.24×10(-11) mol L(-1), respectively. An in vivo subcutaneous xenograft model of CD20-positive B cell lymphoma in BALB/c (nu/nu) mice was also utilized. Drugs were given intravenously on day 14 and 21 after tumor transplantation. In terms of maximal tolerated doses, scFv-LDP-AE at 0.3 mg kg(-1) suppressed tumor growth by 79.3%, and LDM at 0.05 mg kg(-1) by 68.6% (P<0.05). Results suggested scFv-LDP-AE could be a potential candidate for tumor-targeting therapy. PMID:21416325

  11. 替代失活法构建变形链球菌LuxS基因缺陷株%Construction of Streptococcus mutans LuxS gene deletion mutants by long flanking homology polymerase chain reaction

    Institute of Scientific and Technical Information of China (English)

    李月恒; 陈惠珍; 周智; 陈娇; 李锐; 尹一兵; 张雪梅

    2011-01-01

    目的 LuxS基因是变形链球菌生物膜早期形成过程中的关键基因,构建该基因的缺陷菌.方法 采用长臂同源多聚酶链反应(LFH-PCR)方法构建含红霉素耐药基因片段的LuxS基因上、下游同源序列的连接片段,转化到变形链球菌中,在红霉素的平板上筛选缺陷菌株,并采用PCR鉴定.结果 对变形链球菌LuxS基因缺陷菌株进行PCR和DNA序列测定分析证实构建成功.结论 成功构建出变形链球菌LuxS基因的缺陷菌株,为后期针对变形链球菌LuxS基因的相关研究奠定基础.%Objective To construct LuxS deletion mutant of Streptococcus mutans and get the capsule deficient strain. Method Long flanking homology polymerase chain reaction(LFH-PCR) was introduced to generate a gene disruption construct consisting of Emr cassette with long flanking homology regions to the target gene. The electroporation competence of Streptococcus mutans was then transformed with this PCR product. Then the positive transformants were counted on selective agar containing erythromycin and identified by PCR. Result Identification by PCR and sequencing confirmed the validity of the LuxS deletion mutant of Streptococcus mutans. Conclusion The successful construction of the LuxS deletion mutant can be used in further functional genome research.

  12. SPECT imaging of peripheral amyloid in mice by targeting hyper-sulfated heparan sulfate proteoglycans with specific scFv antibodies

    International Nuclear Information System (INIS)

    Introduction: Amyloid deposits are associated with a broad spectrum of disorders including monoclonal gammopathies, chronic inflammation, and Alzheimer's disease. In all cases, the amyloid pathology contains, in addition to protein fibrils, a plethora of associated molecules, including high concentrations of heparan sulfate proteoglycans (HSPGs). Methods: We have evaluated radioiodinated scFvs that bind HS for their ability to image amyloid deposits in vivo. scFv's with different binding characteristics were isolated by phage display using HS extracted from bovine kidney or mouse and human skeletal muscle glycosaminoglycans (GAGs). Following purification and radioiodination, the biodistribution of 125I-scFv's was assessed in mice with inflammation-associated AA amyloidosis or in amyloid-free mice by using SPECT imaging, biodistribution measurements and tissue autoradiography. Results: Four different scFv's all showed binding in vivo to amyloid in the spleen, liver and kidney of diseased mice; however, three of the scFv's also bound to sites within these organs in disease free mice. One scFv specific for hypersulfated HSPGs preferentially bound amyloid and did not accumulate in healthy tissues. Conclusions: These data indicate that HS expressed in amyloid deposits has unique qualities that can be distinguished from HS in normal tissues. A scFv specific for rare hypersulfated HS was used to selectively image AA amyloid in mice with minimal retention in normal tissue.

  13. 建筑供应链库存控制模式与优化方法研究--基于CPFR理论%On the Control Models and Optimization Methods of Construction Supply Chain:Based on the Theory of CPFR

    Institute of Scientific and Technical Information of China (English)

    杜泽超; 王俊

    2015-01-01

    For a long time, inventory management is the difficult point of the supply chain management of construction. Based on the theory of CPFR, this paper studies the inventory control models and optimization methods of construction supply chain and gets the final conclusion.%一直以来,库存管理就是建筑供应链管理的重难点问题。本文基于CPFR理论研究了建筑供应链库存控制模式与优化方法,并得出了最终结论。

  14. CONSTRUCTION AND CHARACTERIZATION OF MUTANT PROINSULIN WITH SHIFTED INTRA-A CHAIN DISULFIDE-BOND%(A11,A12)胰岛素原移位突变体的表征研究

    Institute of Scientific and Technical Information of China (English)

    井健

    2009-01-01

    With PCR technique a mutant proinsulin gene with CysA11Ser and SerA12Cys was constructed to investigate intra A-chain disulfide effect on insulin biological activity. After expression, unfolding and refolding, mutant proinsulin was purified with (A6, A12) intra-A chain disulfide-bond. The mutant proinsulin showed 55% radioimmunoactivity (RIA) and 11% of receptor binding activity of native proinsulin. Mass spectrometry analysis, IEF and amino acid composition assay were carried out to determine mutant proinsulin. Data showed that the A6-A12 intra-A chain disulfide bond was formed, having some negative effect on native proinsulin structure, especially in regard to the stability of insulin receptor binding region.%将(CysA11Ser,SerA12Cys)人胰岛素原移位突变体基因克隆至高效表达质粒pET22b多克隆位点区,构建重组表达质粒pET-A112,并在大肠杆菌E.coli BL21(DE3)pLys中进行了表达.表达产物存在于包涵体中,通过包涵体的分离及二硫键变复性,获得A6-A12二硫键突变型的胰岛素原突变体.SDS-PAGE电泳显示突变型胰岛素原的电泳迁移率与野生型胰岛素原基本相同.质谱检测表明,突变性胰岛素原的分子结构均一、纯度良好,氨基酸组成与理论值基本一致.以野生型人胰岛素原为对照进行的胰岛素受体试验及胰岛素放射免疫试验表明,A6-A12二硫键错接型胰岛素原突变体的受体结合活性和放免活性是野生型胰岛素原的11%与55%.

  15. Functional expression of a single-chain antibody to ErbB-2 in plants and cell-free systems

    Directory of Open Access Journals (Sweden)

    Benevolo Maria

    2006-09-01

    Full Text Available Abstract Background Aberrant signaling by ErbB-2 (HER 2, Neu, a member of the human Epidermal Growth Factor (EGF receptor family, is associated with an aggressive clinical behaviour of carcinomas, particularly breast tumors. Antibodies targeting the ErbB-2 pathway are a preferred therapeutic option for patients with advanced breast cancer, but a worldwide deficit in the manufacturing capacities of mammalian cell bioreactors is foreseen. Methods Herein, we describe a multi-platform approach for the production of recombinant Single chain Fragments of antibody variable regions (ScFvs to ErbB-2 that involves their functional expression in (a bacteria, (b transient as well as stable transgenic tobacco plants, and (c a newly developed cell-free transcription-translation system. Results An ScFv (ScFv800E6 was selected by cloning immunoglobulin sequences from murine hybridomas, and was expressed and fully functional in all the expression platforms, thereby representing the first ScFv to ErbB-2 produced in hosts other than bacteria and yeast. ScFv800E6 was optimized with respect to redox synthesis conditions. Different tags were introduced flanking the ScFv800E6 backbone, with and without spacer arms, including a novel Strep II tag that outperforms conventional streptavidin-based detection systems. ScFv800E6 was resistant to standard chemical radiolabeling procedures (i.e. Chloramine T, displayed a binding ability extremely similar to that of the parental monovalent Fab' fragment, as well as a flow cytometry performance and an equilibrium binding affinity (Ka approximately 2 × 108 M-1 only slightly lower than those of the parental bivalent antibody, suggesting that its binding site is conserved as compared to that of the parental antibody molecule. ScFv800E6 was found to be compatible with routine reagents for immunohistochemical staining. Conclusion ScFv800E6 is a useful reagent for in vitro biochemical and immunodiagnostic applications in oncology

  16. Chain Teleportation

    OpenAIRE

    Lee, Chien-er

    2004-01-01

    By means of the idea of measurements on the crossed space-time nonlocal observables, we extend the mechanism for the two-way quantum teleportation to the chain teleportation among N spatially separated spin-1/2 systems. Since in the process only the local interactions are used, the microcausality is automatically satisfied.

  17. AGING PROCESS OPTIMIZATION FOR A HIGH STRENGTH AND TOUGHNESS OF FV520B MARTENSITIC STEEL%高强高韧FV520B马氏体钢的时效工艺优化

    Institute of Scientific and Technical Information of China (English)

    周倩青; 翟玉春

    2009-01-01

    Low carbon martensitic precipitation hardening stainless steels are widely utilized in many engineering applications due to their high strength with reasonable toughness, ductility and corrosion resistance. However, those properties and their combinations are not always satisfactory to their users. For further improvement of the mechanical properties of these types of steels, a fundamen-tal understanding of the detailed microstructural features with various aging conditions is necessary. Therefore, the effects of aging temperature, aging time and cooling rate on the microstructure and me-chanical properties of a martensitic precipitation hardening stainless steel FV520B were investigated by OM, SEM, TEM and XRD methods. The results show that the steel aged at 630 ℃ for a short time and then furnace cooled, in which a typical lath martensitic with the proper amounts of reverse austenite and fined dispersed precipitates was observed, has a good combination of high strength and high toughness. It could be an optimized aging process for FV520B steel.%通过在600,630,680,700和720℃保温1 h空冷,以及在630℃短时时效后炉冷和空冷的热处理,研究了不同时效温度,时效时间以及冷却方式对沉淀硬化马氏体不锈钢FV520B的组织和力学性能的影响.结果表明,630℃短时时效后钢中即可析出一定量的逆转变奥氏体,且钢中的析出相尺寸较小并弥散分布,由此提出了一种沉淀强化马氏体不锈钢的热处理工艺优化,即FV520B钢经630℃短时时效并炉冷后,可以获得较佳的高强度和高韧性组合.

  18. Single-Chain Fragment Variable Passive Immunotherapies for Neurodegenerative Diseases

    Directory of Open Access Journals (Sweden)

    Liang Huang

    2013-09-01

    Full Text Available Accumulation of misfolded proteins has been implicated in a variety of neurodegenerative diseases including prion diseases, Alzheimer’s disease (AD, Parkinson’s disease (PD, and Huntington’s disease (HD. In the past decade, single-chain fragment variable (scFv -based immunotherapies have been developed to target abnormal proteins or various forms of protein aggregates including Aβ, SNCA, Htt, and PrP proteins. The scFvs are produced by fusing the variable regions of the antibody heavy and light chains, creating a much smaller protein with unaltered specificity. Because of its small size and relative ease of production, scFvs are promising diagnostic and therapeutic reagents for protein misfolded diseases. Studies have demonstrated the efficacy and safety of scFvs in preventing amyloid protein aggregation in preclinical models. Herein, we discuss recent developments of these immunotherapeutics. We review efforts of our group and others using scFv in neurodegenerative disease models. We illustrate the advantages of scFvs, including engineering to enhance misfolded conformer specificity and subcellular targeting to optimize therapeutic action.

  19. Studies on soil to grass transfer factor (Fv) and grass to milk transfer coefficient (Fm) for cesium in Kaiga region

    International Nuclear Information System (INIS)

    Detailed studies were carried out to establish site-specific soil to grass transfer factors (Fv) and grass to cow milk transfer coefficients (Fm) for radioactive cesium (137Cs) and stable cesium (Cs) for Kaiga region, where a nuclear power station has been in operation for more than 10 years. The study included adopted cows, cows of local farmers, and cows from the dairy farm. A grass field was developed specifically for the study and 2 local breed cows were adopted and allowed to graze in this grass field. The soil and grass samples were collected regularly from this field and analyzed for the concentrations of 137Cs and stable Cs to evaluate the soil to grass Fv values. The milk samples from the adopted cows were analyzed for the 137Cs and stable Cs concentrations to evaluate Fm values. For comparison, studies were also carried out in dominant grazing areas in different villages around the nuclear power plant and the cows of local farmers which graze in these areas were identified and milk samples were collected and analyzed regularly. The geometric mean values of Fv were found to be 1.1 × 10−1 and 1.8 × 10−1 for 137Cs and stable Cs, respectively. The Fm of 137Cs had geometric mean values of 1.9 × 10−2 d L−1 and 4.6 × 10−2 d L−1, respectively, for adopted Cows 1 and 2; 1.7 × 10−2 d L−1 for the cows of local farmers, and 4.0 × 10−3 d L−1 for the dairy farm cows. The geometric mean values of Fm for stable Cs were similar to those of 137Cs. The Fm value for the dairy farm cows was an order of magnitude lower than those for local breed cows. The Fm values observed for the local breed cows were also an order of magnitude higher when compared to the many values reported in the literature and in the IAEA publication. Possible reasons for this higher Fm values were identified. The correlation between Fv and Fm values for 137Cs and stable Cs and their dependence on the potassium content (40K and stable K) in the soil and grass were also studied

  20. Optimisation of recombinant protein production in Pichia pastoris:single-chain antibody fragment model protein

    OpenAIRE

    Khatri, N. K. (Narendar Kumar)

    2011-01-01

    Abstract Potential lethal diarrhoea caused by enterotoxigenic Escherichia coli strains is one of the most common diseases in young pigs. It can be cured by single-chain antibody fragments (scFv), which can be produced in recombinant microorganisms. Pichia pastoris, a methylotrophic yeast, is generally considered an interesting production system candidate, as it can secrete properly folded proteins. These proteins accumulate in high concentrations during fermentation, reducing the cost for...

  1. Kinetic Characterisation of a Single Chain Antibody against the Hormone Abscisic Acid: Comparison with Its Parental Monoclonal

    Science.gov (United States)

    Badescu, George O.; Marsh, Andrew; Smith, Timothy R.; Thompson, Andrew J.; Napier, Richard M.

    2016-01-01

    A single-chain Fv fragment antibody (scFv) specific for the plant hormone abscisic acid (ABA) has been expressed in the bacterium Escherichia coli as a fusion protein. The kinetics of ABA binding have been measured using surface plasmon resonance spectrometry (BIAcore 2000) using surface and solution assays. Care was taken to calculate the concentration of active protein in each sample using initial rate measurements under conditions of partial mass transport limitation. The fusion product, parental monoclonal antibody and the free scFv all have low nanomolar affinity constants, but there is a lower dissociation rate constant for the parental monoclonal resulting in a three-fold greater affinity. Analogue specificity was tested and structure-activity binding preferences measured. The biologically-active (+)-ABA enantiomer is recognised with an affinity three orders of magnitude higher than the inactive (-)-ABA. Metabolites of ABA including phaseic acid, dihydrophaseic acid and deoxy-ABA have affinities over 100-fold lower than that for (+)-ABA. These properties of the scFv make it suitable as a sensor domain in bioreporters specific for the naturally occurring form of ABA. PMID:27023768

  2. What is a Responsible Supply Chain?

    Directory of Open Access Journals (Sweden)

    Terje Ingebrigt Vaaland

    2012-02-01

    Full Text Available The paper introduces the concept of responsible supply chain based on two dimensions, the core processes of a supply chain and the concept of corporate social responsibility (CSR. It is suggested that a responsible supply chain is achieved through manifested core values of the supply chain actors, strategies, and tactics. The paper further discusses the individual supply chain actors’ responsibility in securing a responsible supply chain beyond the actors’ direct control. A conceptual model and a definition of a responsible supply chain are offered. Our model not only provides structure to the extant research but also develops new constructs and relationships that improve the conceptualization of the responsible supply chain. The paper is based on a review of 81 research articles published between 2000 and 2010 in which elements of CSR and supply chain processes are included.

  3. Catch Chain

    OpenAIRE

    Talbert, Robert

    2010-01-01

    Catch Chain is a book of poems that traces the journey of a Corrections Officer who attempts to combat issues of isolation, inhumane treatment of inmates and societal rejection in jails by embarking upon a cross-country road trip. However, the same issues the officer initially wrestled with begin cropping up in different cities, on various highways and in a multitude of states. The excitement and adventure of the open road runs parallel to the recurring imprisonment of the guard's mind.

  4. Improved fluoroquinolone detection in ELISA through engineering of a broad-specific single-chain variable fragment binding simultaneously to 20 fluoroquinolones.

    Science.gov (United States)

    Wen, Kai; Nölke, Greta; Schillberg, Stefan; Wang, Zhanhui; Zhang, Suxia; Wu, Congming; Jiang, Haiyang; Meng, Hui; Shen, Jianzhong

    2012-07-01

    Fluoroquinolones (FQs) are a group of synthetic, broad-spectrum antibacterial agents. Due to its extensive use in animal industry and aquaculture, residues of these antibiotics and the emergence of bacteria resistant to FQs have become a major public health issue. To prepare a generic antibody capable of recognizing nearly all FQs, a single-chain variable fragment (scFv) was generated from the murine hybridoma cells C49H1 producing a FQ-specific monoclonal antibody. This scFv was characterized by indirect competitive enzyme-linked immunosorbent assay (ciELISA), and it showed identical binding properties to parental monoclonal antibody: it was capable of recognizing 17 of 20 targeted FQs below maximum residue limits, except for sarafloxacin (SAR), difloxacin (DIF), and trovafloxacin (TRO) which are highly concerned members in the FQs family. In order to broaden the specificity of this scFv to SAR and its analogues (DIF and TRO), protein homology modeling and antibody-ligands docking analysis were employed to identify the potential key amino acid residues involved in hapten antibody. A mutagenesis phage display library was generated by site directed mutagenesis randomizing five aminoacid residues in the third heavy-chain complementarity determining region. After one round of panning against biotinylated norfloxacin (NOR) and four rounds of panning against biotinylated SAR, scFv variants we screened showed up to 10-fold improved IC(50) against SAR, DIF, and TRO in ciELISA while the specificity against other FQs was fully retained. PMID:22549819

  5. SINGLE CHAIN VARIABLE FRAGMENTS OF ANTIBODIES AGAINST DIPHTHERIA TOXIN B-SUBUNIT ISOLATED FROM PHAGE DISPLAY HUMAN ANTIBODY LIBRARY

    Directory of Open Access Journals (Sweden)

    Oliinyk O. S.

    2014-02-01

    Full Text Available Diphtheria toxin is an exoantigen of Corynebacterium diphtheriae that inhibits protein synthesis and kills sensitive cells. The aim of this study was to obtain human recombinant single-chain variable fragment (scFv antibodies against receptor-binding B subunit of diphtheria toxin. 12 specific clones were selected after three rounds of a phage display naїve (unimmunized human antibody library against recombinant B-subunit. scFv DNA inserts from these 12 clones were digested with MvaI, and 6 unique restriction patterns were found. Single-chain antibodies were expressed in Escherichia coli XL1-blue. The recombinant proteins were characterized by immunoblotting of bacterial extracts and detection with an anti-E-tag antibody. The toxin B-subunit-binding function of the single-chain antibody was shown by ELISA. The affinity constants for different clones were found to be from 106 to 108 М–1. Due to the fact, that these antibody fragments recognized epitopes in the receptor-binding Bsubunit of diphtheria toxin, further studies are interesting to evaluate their toxin neutralization properties and potential for therapeutic applications. Obtained scFv-antibodies can also be used for detection and investigation of biological properties of diphtheria toxin.

  6. Towards Intelligent Supply Chains

    DEFF Research Database (Denmark)

    Siurdyban, Artur; Møller, Charles

    2012-01-01

    applied to the context of organizational processes can increase the success rate of business operations. The framework is created using a set of theoretical based constructs grounded in a discussion across several streams of research including psychology, pedagogy, artificial intelligence, learning...... of deploying inapt operations leading to deterioration of profits. To address this problem, we propose a unified business process design framework based on the paradigm of intelligence. Intelligence allows humans and human-designed systems cope with environmental volatility, and we argue that its principles......, business process management and supply chain management. It outlines a number of system tasks combined in four integrated management perspectives: build, execute, grow and innovate, put forward as business process design propositions for Intelligent Supply Chains....

  7. Extended linear chain compounds

    CERN Document Server

    Linear chain substances span a large cross section of contemporary chemistry ranging from covalent polymers, to organic charge transfer com­ plexes to nonstoichiometric transition metal coordination complexes. Their commonality, which coalesced intense interest in the theoretical and exper­ imental solid state physics/chemistry communities, was based on the obser­ vation that these inorganic and organic polymeric substrates exhibit striking metal-like electrical and optical properties. Exploitation and extension of these systems has led to the systematic study of both the chemistry and physics of highly and poorly conducting linear chain substances. To gain a salient understanding of these complex materials rich in anomalous aniso­ tropic electrical, optical, magnetic, and mechanical properties, the conver­ gence of diverse skills and talents was required. The constructive blending of traditionally segregated disciplines such as synthetic and physical organic, inorganic, and polymer chemistry, crystallog...

  8. The multiple readings of 'metaphor' in the classroom: co-construction of inferential chains As múltiplas leituras da 'metáfora' em sala de aula: co-construção de cadeias inferenciais

    Directory of Open Access Journals (Sweden)

    Mara Sophia Zanotto

    2010-01-01

    Full Text Available This paper is part of a research project whose central aim is to empirically investigate the multiple readings of 'metaphors' in literary texts. The methodology employed is interpretive and the main research technique is the 'Group-Think Aloud'. In this paper, the data discussed were generated by a group of readers engaged in the reading of 'The Pulverized Mountain', a poem by Drummond de Andrade. The analysis focuses on the interpretations of the final verses that, due to incongruities presented, constitute enigmas for the reader to decipher. Analysis has shown that the apparent data chaos and complexity has, in fact, an organization in inferential chains co-constructed by metonymic and metaphoric processes.Este trabalho faz parte do projeto de pesquisa que tem como objetivo central investigar empiricamente as múltiplas leituras de metáforas em textos literários. A metodologia é a interpretativista e a técnica principal de pesquisa é o 'Pensar Alto em Grupo'. Neste trabalho são discutidos os dados de um grupo de leitores lendo o poema 'A Montanha Pulverizada', de Drummond de Andrade. O foco da análise são as interpretações dos versos finais, que, devido às incongruências que apresentam, constituem enigmas para o leitor decifrar. A análise mostrou que o aparente caos e complexidade dos dados têm, de fato, uma organização em cadeias inferenciais co-construídas por processos metonímicos e metafóricos.

  9. Stationary Probability Vectors of Higher-order Markov Chains

    OpenAIRE

    Li, Chi-Kwong; Zhang, Shixiao

    2013-01-01

    We consider the higher-order Markov Chain, and characterize the second order Markov chains admitting every probability distribution vector as a stationary vector. The result is used to construct Markov chains of higher-order with the same property. We also study conditions under which the set of stationary vectors of the Markov chain has a certain affine dimension.

  10. Markov Chain Approximations to Singular Stable-like Processes

    OpenAIRE

    Xu, Fangjun

    2012-01-01

    We consider the Markov chain approximations for singular stable-like processes. First we obtain properties of some Markov chains. Then we construct the approximating Markov chains and give a necessary condition for weak convergence of these chains to singular stable-like processes.

  11. Screening and immunological identification of the human ScFv antibody against PSMA%全人源抗PSMA单链抗体的筛选及免疫活性鉴定

    Institute of Scientific and Technical Information of China (English)

    张才田; 刘金霞

    2011-01-01

    Objective To screen and identify a human single-chain variable fragment(scFv) antibody against prostate specific membrane antigen(PSMA) from a human scFv antibody library.Methods Using a synthetic PSMA peptide as the coating antigen, the antibody library was screened by five rounds of combining-eluting-amplification.The phage antibody against PSMA with high specificity was screened out from the human scFv antibody library and its binding ability to the antigen was tested by ELISA.The soluble antibody was produced by plasmids extracted from highly specific clones, whose binding ability to PSMA was further identified by Western blot and immunohistochemistry.The affinity constant of the soluble antibody was measured by non-competitive ELISA.Results The screened phage antibody was specific for PSMA by ELISA.The soluble antibody was also specific for PSMA, its molecular weight was about 30 kD by SDS-PAGE and its affinity constant was about 5.077 × 106 L/mol.Conclusions The screened scFv antibody is specific and has low immunogenicity.It can be further used in the target treatment of malignant tumors.%目的 从全人源单链噬菌体抗体库中筛选出抗前列腺特异性膜抗原(PSMA)特异性单链抗体并进行免疫活性鉴定.方法 以合成的PSMA多肽为抗原,经过五轮吸附-洗脱-扩增,从单链噬菌体抗体库中筛选出特异性抗PSMA噬菌体抗体,ELISA检测其抗原结合能力,并对特异性较强的克隆提取质粒,表达可溶性抗体.Western Blotting和免疫组织化学检测其抗原结合性,非竞争ELSIA法检测其亲和常数.结果 从单链噬菌体抗体库中筛选出的噬菌体抗体,经ELISA鉴定为抗PSMA的特异性噬菌体抗体.抗PSMA可溶性抗体相对分子质量约为3.0×104,与PSMA特异性结合,其亲和常数约为5.077×106L/mol.结论 所得全人源抗PSMA单链抗体保留完整抗体分子结合抗原的特异性,免疫原性弱,是肿瘤导向治疗的理想栽体.

  12. Functional characterization of two scFv-Fc antibodies from an HIV controller selected on soluble HIV-1 Env complexes: a neutralizing V3- and a trimer-specific gp41 antibody.

    Directory of Open Access Journals (Sweden)

    Maria Trott

    Full Text Available HIV neutralizing antibodies (nAbs represent an important tool in view of prophylactic and therapeutic applications for HIV-1 infection. Patients chronically infected by HIV-1 represent a valuable source for nAbs. HIV controllers, including long-term non-progressors (LTNP and elite controllers (EC, represent an interesting subgroup in this regard, as here nAbs can develop over time in a rather healthy immune system and in the absence of any therapeutic selection pressure. In this study, we characterized two particular antibodies that were selected as scFv antibody fragments from a phage immune library generated from an LTNP with HIV neutralizing antibodies in his plasma. The phage library was screened on recombinant soluble gp140 envelope (Env proteins. Sequencing the selected peptide inserts revealed two major classes of antibody sequences. Binding analysis of the corresponding scFv-Fc derivatives to various trimeric and monomeric Env constructs as well as to peptide arrays showed that one class, represented by monoclonal antibody (mAb A2, specifically recognizes an epitope localized in the pocket binding domain of the C heptad repeat (CHR in the ectodomain of gp41, but only in the trimeric context. Thus, this antibody represents an interesting tool for trimer identification. MAb A7, representing the second class, binds to structural elements of the third variable loop V3 and neutralizes tier 1 and tier 2 HIV-1 isolates of different subtypes with matching critical amino acids in the linear epitope sequence. In conclusion, HIV controllers are a valuable source for the selection of functionally interesting antibodies that can be selected on soluble gp140 proteins with properties from the native envelope spike.

  13. Development of a single-chain variable fragment-alkaline phosphatase fusion protein and a sensitive direct competitive chemiluminescent enzyme immunoassay for detection of ractopamine in pork

    Energy Technology Data Exchange (ETDEWEB)

    Dong Jiexian; Li Zhenfeng; Lei Hongtao; Sun Yuanming [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Ducancel, Frederic [CEA, iBiTec-S, Service de Pharmacologie et d' Immnoanalyse (SPI), CEA Saclay, F-91191 Gif sur Yvette (France); Xu Zhenlin [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Boulain, Jean-Claude [CEA, iBiTec-S, Service de Pharmacologie et d' Immnoanalyse (SPI), CEA Saclay, F-91191 Gif sur Yvette (France); Yang Jinyi; Shen Yudong [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China); Wang Hong, E-mail: gzwhongd@63.com [Guangdong Provincial Key Laboratory of Food Quality and Safety, South China Agricultural University, Guangzhou 510642 (China)

    2012-07-29

    Graphical abstract: Detection model of dc-CLEIA based on anti-RAC scFv-AP fusion protein. Highlights: Black-Right-Pointing-Pointer The scFv-AP fusion protein against ractopamine (RAC) was produced. Black-Right-Pointing-Pointer A dc-CLEIA for RAC was developed based on the purified scFv-AP fusion protein. Black-Right-Pointing-Pointer The sensitivity of dc-CLEIA was 10 times as sensitive as dc-ELISA for RAC. Black-Right-Pointing-Pointer Recovery tests from pork samples were studied. Black-Right-Pointing-Pointer Good accuracy was obtained. - Abstract: A rapid, sensitive chemiluminescent enzyme immunoassay (CLEIA) for ractopamine (RAC) based on a single-chain variable fragment (scFv)-alkaline phosphatase (AP) fusion protein was developed. The scFv gene was prepared by cloning the heavy- and light-chain variable region genes (V{sub H} and V{sub L}) from hybridoma cell line AC2, which secretes antibodies against RAC, and assembling V{sub H} and V{sub L} genes with a linker by means of splicing overlap extension polymerase chain reaction. The resulting scFv gene was inserted into the expression vector pLIP6/GN containing AP to produce the fusion protein in Escherichia coli strain BL21. The purified scFv-AP fusion protein was used to develop a direct competitive CLEIA (dcCLEIA) protocol for detection of RAC. The average concentration required for 50% inhibition of binding and the limit of detection of the assay were 0.25 {+-} 0.03 and 0.02 {+-} 0.004 ng mL{sup -1}, respectively, and the linear response range extended from 0.05 to 1.45 ng mL{sup -1}. The assay was 10 times as sensitive as the corresponding enzyme-linked immunosorbent assay based on the same fusion protein. Cross-reactivity studies showed that the fusion protein did not cross react with RAC analogs. DcCLEIA was used to analyze RAC spiked pork samples, and the validation was confirmed by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS). The results showed a good correlation between

  14. Development of a single-chain variable fragment-alkaline phosphatase fusion protein and a sensitive direct competitive chemiluminescent enzyme immunoassay for detection of ractopamine in pork

    International Nuclear Information System (INIS)

    Graphical abstract: Detection model of dc-CLEIA based on anti-RAC scFv-AP fusion protein. Highlights: ► The scFv-AP fusion protein against ractopamine (RAC) was produced. ► A dc-CLEIA for RAC was developed based on the purified scFv-AP fusion protein. ► The sensitivity of dc-CLEIA was 10 times as sensitive as dc-ELISA for RAC. ► Recovery tests from pork samples were studied. ► Good accuracy was obtained. - Abstract: A rapid, sensitive chemiluminescent enzyme immunoassay (CLEIA) for ractopamine (RAC) based on a single-chain variable fragment (scFv)-alkaline phosphatase (AP) fusion protein was developed. The scFv gene was prepared by cloning the heavy- and light-chain variable region genes (VH and VL) from hybridoma cell line AC2, which secretes antibodies against RAC, and assembling VH and VL genes with a linker by means of splicing overlap extension polymerase chain reaction. The resulting scFv gene was inserted into the expression vector pLIP6/GN containing AP to produce the fusion protein in Escherichia coli strain BL21. The purified scFv-AP fusion protein was used to develop a direct competitive CLEIA (dcCLEIA) protocol for detection of RAC. The average concentration required for 50% inhibition of binding and the limit of detection of the assay were 0.25 ± 0.03 and 0.02 ± 0.004 ng mL−1, respectively, and the linear response range extended from 0.05 to 1.45 ng mL−1. The assay was 10 times as sensitive as the corresponding enzyme-linked immunosorbent assay based on the same fusion protein. Cross-reactivity studies showed that the fusion protein did not cross react with RAC analogs. DcCLEIA was used to analyze RAC spiked pork samples, and the validation was confirmed by high-performance liquid chromatography–tandem mass spectrometry (HPLC–MS). The results showed a good correlation between the data of dc-CLEIA and HPLC–MS (R2 > 0.99), indicating that the assay was an efficient analytical method for monitoring food safety.

  15. Multiparameter optimization method and enhanced production of secreted recombinant single-chain variable fragment against the HIV-1 P17 protein from Escherichia coli by fed-batch fermentation.

    Science.gov (United States)

    Paopang, Porntip; Kasinrerk, Watchara; Tayapiwatana, Chatchai; Seesuriyachan, Phisit; Butr-Indr, Bordin

    2016-04-01

    The single-chain fragment variable (scFv) was used to produce a completely functional antigen-binding fragment in bacterial systems. The advancements in antibody engineering have simplified the method of producing Fv fragments and made it more efficient and generally relevant. In a previous study, the scFv anti HIV-1 P17 protein was produced by a batch production system, optimized by the sequential simplex optimization method. This study continued that work in order to enhance secreted scFv production by fed-batch cultivation, which supported high volumetric productivity and provided a large amount of scFvs for diagnostic and therapeutic research. The developments in cell culture media and process parameter settings were required to realize the maximum production of cells. This study investigated the combined optimization methods, Plackett-Burman design (PBD) and sequential simplex optimization, with the aim of optimize feed medium. Fed-batch cultivation with an optimal feeding rate was determined. The result demonstrated that a 20-mL/hr feeding rate of the optimized medium can increase cell growth, total protein production, and scFv anti-p17 activity by 4.43, 1.48, and 6.5 times more than batch cultivation, respectively. The combined optimization method demonstrated novel power tools for the optimization strategy of multiparameter experiments. PMID:25831436

  16. A single-chain variable fragment intrabody prevents intracellular polymerization of Z α1-antitrypsin while allowing its antiproteinase activity.

    Science.gov (United States)

    Ordóñez, Adriana; Pérez, Juan; Tan, Lu; Dickens, Jennifer A; Motamedi-Shad, Neda; Irving, James A; Haq, Imran; Ekeowa, Ugo; Marciniak, Stefan J; Miranda, Elena; Lomas, David A

    2015-06-01

    Mutant Z α1-antitrypsin (E342K) accumulates as polymers within the endoplasmic reticulum (ER) of hepatocytes predisposing to liver disease, whereas low levels of circulating Z α1-antitrypsin lead to emphysema by loss of inhibition of neutrophil elastase. The ideal therapy should prevent polymer formation while preserving inhibitory activity. Here we used mAb technology to identify interactors with Z α1-antitrypsin that comply with both requirements. We report the generation of an mAb (4B12) that blocked α1-antitrypsin polymerization in vitro at a 1:1 molar ratio, causing a small increase of the stoichiometry of inhibition for neutrophil elastase. A single-chain variable fragment (scFv) intrabody was generated based on the sequence of mAb4B12. The expression of scFv4B12 within the ER (scFv4B12KDEL) and along the secretory pathway (scFv4B12) reduced the intracellular polymerization of Z α1-antitrypsin by 60%. The scFv4B12 intrabody also increased the secretion of Z α1-antitrypsin that retained inhibitory activity against neutrophil elastase. MAb4B12 recognized a discontinuous epitope probably located in the region of helices A/C/G/H/I and seems to act by altering protein dynamics rather than binding preferentially to the native state. This novel approach could reveal new target sites for small-molecule intervention that may block the transition to aberrant polymers without compromising the inhibitory activity of Z α1-antitrypsin. PMID:25757566

  17. Targeting Prostate Cancer Cells In Vivo Using a Rapidly Internalizing Novel Human Single-Chain Antibody Fragment

    Science.gov (United States)

    He, Jiang; Wang, Yong; Feng, Jinjin; Zhu, Xiaodong; Lan, Xiaoli; Iyer, Arun K.; Zhang, Niu; Seo, Youngho; VanBrocklin, Henry F.; Liu, Bin

    2010-01-01

    Human antibodies targeting prostate cancer cell surface epitopes may be useful for imaging and therapy. The objective of this study was to evaluate the tumor targeting of an internalizing human antibody fragment, a small-size platform, to provide high contrast in a mouse model of human prostate carcinoma. Methods A prostate tumor-targeting single-chain antibody fragment (scFv), UA20, along with a nonbinding control scFv, N3M2, were labeled with 99mTc and evaluated for binding and rapid internalization into human prostate tumor cells in vitro and tumor homing in vivo using xenograft models. For the in vitro studies, the labeled UA20 scFv was incubated at 37°C for 1 h with metastatic prostate cancer cells (DU145) to assess the total cellular uptake versus intracellular uptake. For the animal studies, labeled UA20 and N3M2 scFvs were administered to athymic mice implanted subcutaneously with DU145 cells. Mice were imaged with small-animal SPECT/CT with concomitant biodistribution at 1 and 3 h after injection. Results The UA20 scFv was labeled in 55%–65% yield and remained stable in phosphate buffer within 24 h. The labeled UA20 scFv was taken up specifically by prostate tumor cells. Internalization was rapid, because incubation at 37°C for less than 1 h resulted in 93% internalization of total cell-associated scFvs. In animal studies, SPECT/CT showed significant tumor uptake as early as 1 h after injection. At 3 h after injection, tumor uptake was 4.4 percentage injected dose per gram (%ID/g), significantly greater than all organs or tissues studied (liver, 2.7 %ID/g; other organs or tissues, <1 %ID/g), except the kidneys (81.4 %ID/g), giving tumor-to-blood and tumor-to-muscle ratios of 12:1 and 70:1, respectively. In contrast, the control antibody exhibited a tumor uptake of only 0.26 %ID/g, similar to that of muscle and fat. Tumor-specific targeting was evidenced by reduced tumor uptake of nearly 70% on administration of a 10-fold excess of unlabeled UA20 scFv

  18. 基于港口服务供应链的高职港口物流专业群建设实践%Practice of Construction of Higher Vocational Port Logistics Specialty Group Based on Port Service Supply Chain

    Institute of Scientific and Technical Information of China (English)

    靳荣利; 王贵斌; 陈艳玲

    2014-01-01

    阐述了基于港口服务供应链构建港口物流专业群的基本内涵,并对基于港口服务供应链的高职港口物流专业群建设的实践进行了探讨。%In this paper, we proposed the basic connotation of the construction of the port logistics specialty group based on the port service supply chain and introduced the practice of the construction of the higher vocational port logistics specialty group.

  19. Remote monitoring of solar PV system for rural areas using GSM, V-F & F-V converters

    Science.gov (United States)

    Tejwani, R.; Kumar, G.; Solanki, C. S.

    2016-05-01

    The Small capacity photovoltaic (PV) systems like solar lantern and home lighting systems installed in remote rural area often fail without any prior warning due to lack of monitoring and maintenance. This paper describes implementation of remote monitoring for small capacity solar PV system that uses GSM voice channel for communication. Through GSM analog signal of sine wave with frequency range 300–3500 Hz and amplitude range 2.5–4 V is transmitted. Receiver is designed to work in the same frequency range. The voltage from solar PV system in range of 2 to 7.5 V can be converted to frequency directly at the transmitting end. The frequency range from 300–6000 Hz can be sensed and directly converted to voltage signal at receiving end. Testing of transmission and reception of analog signal through GSM voice channel is done for voltage to frequency (V-F) and frequency to voltage (F-V) conversions.

  20. Antecedents and Dimensions of Supply Chain Robustness

    OpenAIRE

    Durach, Christian F.; Wieland, Andreas; Machuca, Jose A.D.

    2015-01-01

    Purpose – The purpose of this paper is to provide groundwork for an emerging theory of supply chain robustness – which has been conceptualized as a dimension of supply chain resilience – through reviewing and synthesizing related yet disconnected studies. The paper develops a formal definition of supply chain robustness to build a framework that captures the dimensions, antecedents and moderators of the construct as discussed in the literature. Design/methodology/approach – The...

  1. High throughput ranking of recombinant avian scFv antibody fragments from crude lysates using the Biacore A100.

    Science.gov (United States)

    Leonard, Paul; Säfsten, Pär; Hearty, Stephen; McDonnell, Barry; Finlay, William; O'Kennedy, Richard

    2007-06-30

    Advances in molecular evolution strategies have made it possible to identify antibodies with exquisite specificities and also to fine-tune their biophysical properties for practically any specified application. Depending on the desired function, antibody/antigen interactions can be long-lived or short-lived and, therefore, particular attention is needed when seeking to identify antibodies with specific reaction-rate and affinity properties. Surface plasmon resonance (SPR) biosensors routinely generate sensitive and reliable kinetic data from antibody/antigen interactions for both therapeutic and diagnostic applications. However, many kinetic-based screening assays require rigorous sample preparation and purification prior to analysis. To ameliorate this problem, we developed a rapid and reliable assay for characterising recombinant scFv antibody fragments, directly from crude bacterial lysates. Ninety-six scFv antibodies derived from chickens immunised with C-reactive protein (CRP) were selected by phage display and evaluated using the Biacore A100 protein interaction array system. Antibodies were captured from crude bacterial extracts on the sensor chip surface and ranked based on the percentage of the complex left (% left) after dissociation in buffer. Kinetic rate constants (k(a) and k(d)) and affinity (K(D)) data were obtained for six clones that bound monomeric CRP across a broad affinity range (2.54 x 10(-8) to 3.53 x 10(-10) M). Using this assay format the A100 biosensor yielded high quality kinetic data, permitting the screening of nearly 400 antibody clones per day. PMID:17532001

  2. Affinity Purification of Tumor Necrosis Factor-α Expressed in Raji Cells by Produced scFv Antibody Coupled CNBr-Activated Sepharose

    Directory of Open Access Journals (Sweden)

    Safar Farajnia

    2013-02-01

    Full Text Available Purpose: Recombinant tumor necrosis factor-alpha (TNF-α has been utilized as an antineoplastic agent for the treatment of patients with melanoma and sarcoma. It targets tumor cell antigens by impressing tumor-associated vessels. Protein purification with affinity chromatography has been widely used in the downstream processing of pharmaceutical-grade proteins. Methods: In this study, we examined the potential of our produced anti-TNF-scFv fragments for purification of TNF-α produced by Raji cells. he Raji cells were induced by lipopolysaccharides (LPS to express TNF-α. Western blotting and Fluorescence-activated cell sorting (FACS flow cytometry analyses were used to evaluate the TNF-α expression. The anti-TNF-α scFv selected from antibody phage display library was coupled to CNBr-activated sepharose 4B beads used for affinity purification of expressed TNF-α and the purity of the protein was assessed by SDS-PAGE. Results: Western blot and FACS flow cytometry analyses showed the successful expression of TNF-α with Raji cells. SDS-PAGE analysis showed the performance of scFv for purification of TNF-α protein with purity over 95%. Conclusion: These findings confirm not only the potential of the produced scFv antibody fragments but also this highly pure recombinant TNF-α protein can be applied for various in vitro and in vivo applications.

  3. FvBck1, a component of cell wall integrity MAP kinase pathway, is required for virulence and oxidative stress response in sugarcane Pokkah Boeng pathogen.

    Science.gov (United States)

    Zhang, Chengkang; Wang, Jianqiang; Tao, Hong; Dang, Xie; Wang, Yang; Chen, Miaoping; Zhai, Zhenzhen; Yu, Wenying; Xu, Liping; Shim, Won-Bo; Lu, Guodong; Wang, Zonghua

    2015-01-01

    Fusarium verticillioides (formerly F. moniliforme) is suggested as one of the causal agents of Pokkah Boeng, a serious disease of sugarcane worldwide. Currently, detailed molecular and physiological mechanism of pathogenesis is unknown. In this study, we focused on cell wall integrity MAPK pathway as one of the potential signaling mechanisms associated with Pokkah Boeng pathogenesis. We identified FvBCK1 gene that encodes a MAP kinase kinase kinase homolog and determined that it is not only required for growth, micro- and macro-conidia production, and cell wall integrity but also for response to osmotic and oxidative stresses. The deletion of FvBCK1 caused a significant reduction in virulence and FB1 production, a possibly carcinogenic mycotoxin produced by the fungus. Moreover, we found the expression levels of three genes, which are known to be involved in superoxide scavenging, were down regulated in the mutant. We hypothesized that the loss of superoxide scavenging capacity was one of the reasons for reduced virulence, but overexpression of catalase or peroxidase gene failed to restore the virulence defect in the deletion mutant. When we introduced Magnaporthe oryzae MCK1 into the FvBck1 deletion mutant, while certain phenotypes were restored, the complemented strain failed to gain full virulence. In summary, FvBck1 plays a diverse role in F. verticillioides, and detailed investigation of downstream signaling pathways will lead to a better understanding of how this MAPK pathway regulates Pokkah Boeng on sugarcane. PMID:26500635

  4. FvBck1, a Component of Cell Wall Integrity MAP Kinase Pathway, is Required for Virulence and Oxidative Stress Response in Sugarcane Pokkah Boeng Pathogen

    Directory of Open Access Journals (Sweden)

    Chengkang eZhang

    2015-10-01

    Full Text Available Fusarium verticillioides (formerly F. moniliforme is suggested as one of the causal agents of Pokkah Boeng, a serious disease of sugarcane worldwide. Currently, detailed molecular and physiological mechanism of pathogenesis is unknown. In this study, we focused on cell wall integrity MAPK pathway as one of the potential signaling mechanisms associated with Pokkah Boeng pathogenesis. We identified FvBCK1 gene that encodes a MAP kinase kinase kinase homolog and determined that it is not only required for growth, micro- and macro-conidia production, and cell wall integrity but also for response to osmotic and oxidative stresses. The deletion of FvBCK1 caused a significant reduction in virulence and FB1 production, a carcinogenic mycotoxin produced by the fungus. Moreover, we found the expression levels of three genes, which are known to be involved in superoxide scavenging, were down regulated in the mutant. We hypothesized that the loss of superoxide scavenging capacity was one of the reasons for reduced virulence, but overexpression of catalase or peroxidase gene failed to restore the virulence defect in the deletion mutant. When we introduced Magnaporthe oryzae MCK1 into the FvBck1 deletion mutant, while certain phenotypes were restored, the complemented strain failed to gain full virulence. In summary, FvBck1 plays a diverse role in F. verticillioides, and detailed investigation of downstream signaling pathways will lead to a better understanding of how this MAPK pathway regulates Pokkah Boeng on sugarcane.

  5. Generation of anti-idiotype scFv for pharmacokinetic measurement in lymphoma patients treated with chimera anti-CD22 antibody SM03.

    Directory of Open Access Journals (Sweden)

    Qi Zhao

    Full Text Available Pre-clinical and clinical studies of therapeutic antibodies require highly specific reagents to examine their immune responses, bio-distributions, immunogenicity, and pharmacodynamics in patients. Selective antigen-mimicking anti-idiotype antibody facilitates the assessment of therapeutic antibody in the detection, quantitation and characterization of antibody immune responses. Using mouse specific degenerate primer pairs and splenocytic RNA, we generated an idiotype antibody-immunized phage-displayed scFv library in which an anti-idiotype antibody against the therapeutic chimera anti-CD22 antibody SM03 was isolated. The anti-idiotype scFv recognized the idiotype of anti-CD22 antibody and inhibited binding of SM03 to CD22 on Raji cell surface. The anti-idiotype scFv was subsequently classified as Ab2γ type. Moreover, our results also demonstrated firstly that the anti-idiotype scFv could be used for pharmacokinetic measurement of circulating residual antibody in lymphoma patients treated with chimera anti-CD22 monoclonal antibody SM03. Of important, the present approach could be easily adopted to generate anti-idiotype antibodies for therapeutic antibodies targeting membrane proteins, saving the cost and time for producing a soluble antigen.

  6. Construction of HLA/Peptide Tetramer with Peptide-Linked β2 Microglobulin

    Institute of Scientific and Technical Information of China (English)

    沈传来; ChienchungCHANG; 张建琼; 郭薇; 孟凡岩; 谢维

    2004-01-01

    Analysis of the frequency of antigen-specific cytotoxic T lymphocytes (CTLs) ex vivo is largely dependent on the use of MHC/peptide tetramers. However, the latter reagents have not been widely available, most likely because of their costly and time-consuming production. In this report we utilized an economic strategy to construct HLA/peptide tetramers with recombinant peptide-linked β2 microglobulin (β2m). The HLA-A2-restricted, melanoma antigen MARTl-derived peptide MARTI27-35 (AAGIGILTV) was fused to the N terminus of human β2m through a 15-amino acid (aa)-long linker before being refolded with the recombinant biotinylated HLA-A2 heavy chain ectodomain. The resulted 2-component (2C) monomer was then tetramerized with phycoerythin-labeled streptavidin. The experimental result showed that the 2C HLA-A2/MARTI27-35 monomer was shown to bind to the HLA class Ⅰ complex-specific monoclonal antibody W6/32 and the HLA-A2/MARTI27-35 complex-specific single chain antibody fragment (scFv) 8.3, suggesting the correctness of its specificity. Furthermore, the 2C HLA-A2/MARTI27-35 tetramer detected a specific CD8+ T cell population in HLA-A2-restricted melanoma infiltrating lymphocytes as the conventional 3C HLA-A2/MARTI27-35 tetramer. The yield of 2C HLA-A2/MARTI27-35 monomerwas 2.5 times more than that of the conventional 3C monomer. Taken together, these data indicate that the HLA-A2/MARTI27-35 tetramer can be generated conveniently through the use of MARTI27-35 peptide-β2m fusion proteins, which can facilitate the monitoring of HLA-A2-restricted, MARTl-specific CTL responses in patients with melanoma.

  7. Chain reaction

    International Nuclear Information System (INIS)

    Chain Reaction is a work of recent American political history. It seeks to explain how and why America came to depend so heavily on its experts after World War II, how those experts translated that authority into political clout, and why that authority and political discretion declined in the 1970s. The author's research into the internal memoranda of the Atomic Energy Commission substantiates his argument in historical detail. It was not the ravages of American anti-intellectualism, as so many scholars have argued, that brought the experts back down to earth. Rather, their decline can be traced to the very roots of their success after World War II. The need to over-state anticipated results in order to garner public support, incessant professional and bureaucratic specialization, and the sheer proliferation of expertise pushed arcane and insulated debates between experts into public forums at the same time that a broad cross section of political participants found it easier to gain access to their own expertise. These tendencies ultimately undermined the political influence of all experts. (author)

  8. Construction of Green Supply Chain Management System and Implementation Steps of China's Phytochemistry Enterprise%中国植提企业绿色供应链管理体系构建与实施步骤研究

    Institute of Scientific and Technical Information of China (English)

    胡雪松; 杜娅

    2011-01-01

    分析了中国植提企业实施绿色供应链管理的必要性,构建了绿色供应链管理职能体系,提出了实施绿色供应链管理的步骤.%The necessity of implementing green supply chain management in China's phytochemistry enterprises is analyzed and a functions system of green supply chain management is built, and the implementation steps are proposed.

  9. The Invariance Principle for p-(→θ) Chain

    Institute of Scientific and Technical Information of China (English)

    Di He HU; Zheng Yan XIAO

    2007-01-01

    There are two parts in this paper. In the first part we construct the Maxkov chain in random environment(MCRE), the skew product Markov chain and p-(→θ) chain from a random transition matrix and a two-dimensional probability distribution, and in the second part we prove that the invariance principle for p-(→θ) chain, a more complex non-homogeneons Markov chain, is true under some reasonable conditions. This result is more powerful.

  10. XTROEM-FV: a new code for computational astrophysics based on very high order finite-volume methods - II. Relativistic hydro- and magnetohydrodynamics

    Science.gov (United States)

    Núñez-de la Rosa, Jonatan; Munz, Claus-Dieter

    2016-07-01

    In this work, we discuss the extension of the XTROEM-FV code to relativistic hydrodynamics and magnetohydrodynamics. XTROEM-FV is a simulation package for computational astrophysics based on very high order finite-volume methods on Cartesian coordinates. Arbitrary spatial high order of accuracy is achieved with a weighted essentially non-oscillatory (WENO) reconstruction operator, and the time evolution is carried out with a strong stability preserving Runge-Kutta scheme. In XTROEM-FV has been implemented a cheap, robust, and accurate shock-capturing strategy for handling complex shock waves problems, typical in an astrophysical environment. The divergence constraint of the magnetic field is tackled with the generalized Lagrange multiplier divergence cleaning approach. Numerical computations of smooth flows for the relativistic hydrodynamics and magnetohydrodynamics equations are performed and confirm the high-order accuracy of the main reconstruction algorithm for such kind of flows. XTROEM-FV has been subject to a comprehensive numerical benchmark, especially for complex flows configurations within an astrophysical context. Computations of problems with shocks with very high order reconstruction operators up to seventh order are reported. For instance, one-dimensional shock tubes problems for relativistic hydrodynamics and magnetohydrodynamics, as well as two-dimensional flows like the relativistic double Mach reflection problem, the interaction of a shock wave with a bubble, the relativistic Orszag-Tang vortex, the cylindrical blast wave problem, the rotor problem, the Kelvin-Helmholtz instability, and an astrophysical slab jet. XTROEM-FV represents a new attempt to simulate astrophysical flow phenomena with very high order numerical methods.

  11. Crater Chains

    Science.gov (United States)

    2003-01-01

    [figure removed for brevity, see original site] The large crater at the top of this THEMIS visible image has several other craters inside of it. Most noticeable are the craters that form a 'chain' on the southern wall of the large crater. These craters are a wonderful example of secondary impacts. They were formed when large blocks of ejecta from an impact crashed back down onto the surface of Mars. Secondaries often form radial patterns around the impact crater that generated them, allowing researchers to trace them back to their origin.Note: this THEMIS visual image has not been radiometrically nor geometrically calibrated for this preliminary release. An empirical correction has been performed to remove instrumental effects. A linear shift has been applied in the cross-track and down-track direction to approximate spacecraft and planetary motion. Fully calibrated and geometrically projected images will be released through the Planetary Data System in accordance with Project policies at a later time.NASA's Jet Propulsion Laboratory manages the 2001 Mars Odyssey mission for NASA's Office of Space Science, Washington, D.C. The Thermal Emission Imaging System (THEMIS) was developed by Arizona State University, Tempe, in collaboration with Raytheon Santa Barbara Remote Sensing. The THEMIS investigation is led by Dr. Philip Christensen at Arizona State University. Lockheed Martin Astronautics, Denver, is the prime contractor for the Odyssey project, and developed and built the orbiter. Mission operations are conducted jointly from Lockheed Martin and from JPL, a division of the California Institute of Technology in Pasadena.Image information: VIS instrument. Latitude 19.3, Longitude 347.5 East (12.5 West). 19 meter/pixel resolution.

  12. Graphs: Associated Markov Chains

    OpenAIRE

    Murthy, Garimella Rama

    2012-01-01

    In this research paper, weighted / unweighted, directed / undirected graphs are associated with interesting Discrete Time Markov Chains (DTMCs) as well as Continuous Time Markov Chains (CTMCs). The equilibrium / transient behaviour of such Markov chains is studied. Also entropy dynamics (Shannon entropy) of certain structured Markov chains is investigated. Finally certain structured graphs and the associated Markov chains are studied.

  13. Screening for a human single chain Fv antibody against epitope on amyloid-beta 1-40 from a human phage display library

    Institute of Scientific and Technical Information of China (English)

    ZHAO Zhen-fu; GAO Guo-quan; LIU Shu; ZOU Jun-tao; XIE Yao; YUAN Qun-fang; WANG Hua-qiao; YAO Zhi-bin

    2007-01-01

    @@ Amyloid-beta peptides (Aβ) are believed to be responsible for the mental decline in patients with Alzheimer's disease (AD). In 1999, Schenk et al1 reported that immunization with Aβ attenuated AD-like pathology in the PDAPP mouse, and developed a new vaccination approach to AD.

  14. Radiology's value chain.

    Science.gov (United States)

    Enzmann, Dieter R

    2012-04-01

    A diagnostic radiology value chain is constructed to define its main components, all of which are vulnerable to change, because digitization has caused disaggregation of the chain. Some components afford opportunities to improve productivity, some add value, while some face outsourcing to lower labor cost and to information technology substitutes, raising commoditization risks. Digital image information, because it can be competitive at smaller economies of scale, allows faster, differential rates of technological innovation of components, initiating a centralization-to-decentralization technology trend. Digitization, having triggered disaggregation of radiology's professional service model, may soon usher in an information business model. This means moving from a mind-set of "reading images" to an orientation of creating and organizing information for greater accuracy, faster speed, and lower cost in medical decision making. Information businesses view value chain investments differently than do small professional services. In the former model, producing a better business product will extend image interpretation beyond a radiologist's personal fund of knowledge to encompass expanding external imaging databases. A follow-on expansion with integration of image and molecular information into a report will offer new value in medical decision making. Improved interpretation plus new integration will enrich and diversify radiology's key service products, the report and consultation. A more robust, information-rich report derived from a "systems" and "computational" radiology approach will be facilitated by a transition from a professional service to an information business. Under health care reform, radiology will transition its emphasis from volume to greater value. Radiology's future brightens with the adoption of a philosophy of offering information rather than "reads" for decision making. Staunchly defending the status quo via turf wars is unlikely to constitute a

  15. Construction of Customization Marketing Service Model for Agricultural Product Cold Chain Logistics Enterprises%农产品冷链物流企业的定制营销服务模型构建

    Institute of Scientific and Technical Information of China (English)

    原惠群; 舒文定

    2012-01-01

    在借鉴第三方物流企业定制营销模式的基础上,通过介绍农产品冷链物流企业实现定制化营销服务的依据与模式,对农产品冷链物流企业定制化营销中的各个模块进行了具体的说明,为我国农产品冷链物流企业开展满足客户个性化需求的定制营销提供一定的借鉴。%China agricultural cold chain has a vast market,a trend of diversification and personalization for the customer demand.According to the personalized demand of customers,the customization marleting is increasingy respected and widely recognized by enterprises and customers.Therefore,it is particularly important to further analyse customization behavior of agricultural cold chain logistics enterprise.Based on the reference to the customization marketing model of the third party logistics enterprise,this paper describes each module of customization marketing in details,through introducing the basis and mode of achieving customized marketing services for the agricultural product cold chain logistics enterprises.It is hoped that a certain reference can be provided to our country agricultural cold chain logistics enterprises for implementing the customization marketing to meet customer demand for individual requirements.

  16. On Markov Chains and Filtrations

    OpenAIRE

    Spreij, Peter

    1997-01-01

    In this paper we rederive some well known results for continuous time Markov processes that live on a finite state space.Martingale techniques are used throughout the paper. Special attention is paid to the construction of a continuous timeMarkov process, when we start from a discrete time Markov chain. The Markov property here holds with respect tofiltrations that need not be minimal.

  17. The comparison of Virtual Machine Migration Performance between XEN-HVM, XEN-PV, Open-VZ, KVM-FV, KVM-PV

    Directory of Open Access Journals (Sweden)

    Igli Tafa

    2011-08-01

    Full Text Available Based on our previous experience we want to compare theperformance between five hypervisors: XEN-PV, XENHVM,Open-VZ,KVM-FV,KVM-PV We have simulated themigration of a virtual machine by using a warning failureapproach. Based on some experiments we have compared CPUConsumption, Memory Utilization, Total Migration Time andDowntime. We have also tested the hypervisor’s performance bychanging the packet’s size from 1500 byte to 32 byte. From thesetests we have concluded that Open-VZ has a bigger CPUConsumption than XEN-PV, but the Total Migration time issmaller than in XEN-PV. XEN-HVM has a worse performancethan XEN-PV, especially regarding to Downtimeparameter.KVM-FV has the worse performance but it canimprove by modifying it, so in this way we use KVM-PV whichhas approximately the same performance with Xen-HVM

  18. FvBck1, a component of cell wall integrity MAP kinase pathway, is required for virulence and oxidative stress response in sugarcane Pokkah Boeng pathogen

    OpenAIRE

    Zhang, Chengkang; Wang, Jianqiang; Tao, Hong; Dang, Xie; Wang, Yang; Chen, Miaoping; Zhai, Zhenzhen; Yu, Wenying; Xu, Liping; Shim, Won-Bo; Lu, Guodong; Wang, Zonghua

    2015-01-01

    Fusarium verticillioides (formerly F. moniliforme) is suggested as one of the causal agents of Pokkah Boeng, a serious disease of sugarcane worldwide. Currently, detailed molecular and physiological mechanism of pathogenesis is unknown. In this study, we focused on cell wall integrity MAPK pathway as one of the potential signaling mechanisms associated with Pokkah Boeng pathogenesis. We identified FvBCK1 gene that encodes a MAP kinase kinase kinase homolog and determined that it is not only r...

  19. FvBck1, a Component of Cell Wall Integrity MAP Kinase Pathway, is Required for Virulence and Oxidative Stress Response in Sugarcane Pokkah Boeng Pathogen

    OpenAIRE

    Chengkang eZhang; Jianqiang eWang; Hong eTao; Xie eDang; Yang eWang; Miaoping eChen; Zhenzhen eZhai; Wenying eYu; Liping eXu; Won-Bo eShim; Guodong eLu; Zonghua eWang

    2015-01-01

    Fusarium verticillioides (formerly F. moniliforme) is suggested as one of the causal agents of Pokkah Boeng, a serious disease of sugarcane worldwide. Currently, detailed molecular and physiological mechanism of pathogenesis is unknown. In this study, we focused on cell wall integrity MAPK pathway as one of the potential signaling mechanisms associated with Pokkah Boeng pathogenesis. We identified FvBCK1 gene that encodes a MAP kinase kinase kinase homolog and determined that it is not only r...

  20. Ophiophagus hannah Venom: Proteome, Components Bound by Naja kaouthia Antivenin and Neutralization by N. kaouthia Neurotoxin-Specific Human ScFv

    Directory of Open Access Journals (Sweden)

    Witchuda Danpaiboon

    2014-05-01

    Full Text Available Venomous snakebites are an important health problem in tropical and subtropical countries. King cobra (Ophiophagus hannah is the largest venomous snake found in South and Southeast Asia. In this study, the O. hannah venom proteome and the venom components cross-reactive to N. kaouthia monospecific antivenin were studied. O. hannah venom consisted of 14 different protein families, including three finger toxins, phospholipases, cysteine-rich secretory proteins, cobra venom factor, muscarinic toxin, L-amino acid oxidase, hypothetical proteins, low cysteine protein, phosphodiesterase, proteases, vespryn toxin, Kunitz, growth factor activators and others (coagulation factor, endonuclease, 5’-nucleotidase. N. kaouthia antivenin recognized several functionally different O. hannah venom proteins and mediated paratherapeutic efficacy by rescuing the O. hannah envenomed mice from lethality. An engineered human ScFv specific to N. kaouthia long neurotoxin (NkLN-HuScFv cross-neutralized the O. hannah venom and extricated the O. hannah envenomed mice from death in a dose escalation manner. Homology modeling and molecular docking revealed that NkLN-HuScFv interacted with residues in loops 2 and 3 of the neurotoxins of both snake species, which are important for neuronal acetylcholine receptor binding. The data of this study are useful for snakebite treatment when and where the polyspecific antivenin is not available. Because the supply of horse-derived antivenin is limited and the preparation may cause some adverse effects in recipients, a cocktail of recombinant human ScFvs for various toxic venom components shared by different venomous snakes, exemplified by the in vitro produced NkLN-HuScFv in this study, should contribute to a possible future route for an improved alternative to the antivenins.

  1. Health supply chain management.

    Science.gov (United States)

    Zimmerman, Rolf; Gallagher, Pat

    2010-01-01

    This chapter gives an educational overview of: * The actual application of supply chain practice and disciplines required for service delivery improvement within the current health environment. * A rationale for the application of Supply Chain Management (SCM) approaches to the Health sector. * The tools and methods available for supply chain analysis and benchmarking. * Key supply chain success factors. PMID:20407173

  2. Silicone chain extender

    DEFF Research Database (Denmark)

    2015-01-01

    The present invention relates to a silicone chain extender, more particularly a chain extender for silicone polymers and copolymers, to a chain extended silicone polymer or copolymer and to a functionalized chain extended silicone polymer or copolymer, to a method for the preparation thereof and...

  3. Profile data from CTD casts aboard the F/V Ocean Explorer in the Arctic Ocean and Beaufort Sea from 2008-08-06 to 2008-08-22 (NODC Accession 0001920)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This profile data aboard the F/V Ocean Explorer in the Arctic Ocean and Beaufort Sea from August 6, 2008 to August 22, 2008 was supported by the Minerals Management...

  4. Activation of erythropoietin receptors by Friend viral gp55 and by erythropoietin and down-modulation by the murine Fv-2r resistance gene

    International Nuclear Information System (INIS)

    The leukemogenic membrane glycoprotein (gp55) encoded by Friend spleen focus-forming virus appears to bind to erythropoietin receptors (EpoR) to stimulate erythroblastosis. To directly compare the effects of gp55 with erythropoietin (Epo), the authors produced retrovirions that encode either gp55, Epo, or EpoR. After infection with EpoR virus, interleukin 3-dependent DA-3 cells bound 125I-labeled Epo and grew without interleukin 3 in the presence of Epo. These latter cells, but not parental DA-3 cells, became factor-independent after superinfection either with Epo virus or with Friend spleen focus-forming virus. In addition, Epo virus caused a disease in mice that mimicked Friend erythroleukemia. Although Fv-2r homozygotes are susceptible to all other retroviral diseases, they are resistant to both Epo viral and Friend viral erythroleukemia. These results indicate that both gp55 and Epo stimulate EpoR and that the Fv-2 gene encodes a protein that controls response to these ligands. However, the Fv-2 protein is not EpoR because the corresponding genes map to opposite ends of mouse chromosome 9. These results have important implications for understanding signal transduction by EpoR and the role of host genetic variation in controlling susceptibility to an oncogenic protein

  5. 5. technical conference 'car body chain': modern concepts in car body engineering - design, construction, materials, projecting, fabrication, quality, recycling; 5. Fachtagung 'Prozesskette Karosserie': Konzepte der Neuzeit im Karosseriebau - Design, Konstruktion, Werkstoffeinsatz, Planung, Fertigung, Qualitaet

    Energy Technology Data Exchange (ETDEWEB)

    Westkaemper, E.; Schraft, R.D. (eds.)

    2000-07-01

    The conference discussed the optimisation of the car body process chain and the integration of new technologies. Cost reduction and lightweight construction with new materials were the key issues. This proceedings volume comprises 19 papers, all of which can be found in this database. [German] Fuer die Tagung 'Konzepte der Neuzeit im Karosseriebau' steht die Aufgabe bevor, die Prozesskette zu optimieren und zu beschleunigen sowie neue Technologien einzubinden. Die wichtigsten Ziele, die Senkung der Herstellungskosten und die Reduzierung der Fahrzeugmasse durch den Einsatz von innovativen Werkstoffen, werden an diesen Objekten dargestellt. In dieser Tagung sind 19 Referate abgehalten und befinden sich in dem vorliegenden Band. (orig.)

  6. Risk Migration In Supply Chain Inventory Financing Service

    OpenAIRE

    Zheng Qin; Xiaochao Ding

    2011-01-01

    Inventory financing affects the risks of both for banks and supply chain companies. Traditionally, supply chain research focus more on material flow than financial. We construct a supply chain financing risk-information migration model (RMM). In this model, we discussed the preconditions to adopt inventory financing when the enterprises are facing cash constraints. And we simulated the whole operate of supply chain and bank behavior with Matlab. The simulation result shows if loan conditions ...

  7. Evaluation and characterization of 30 solar home systems (SHS) Ovonic-Unisolar; Caracterizacion y evaluacion de 30 sistemas FV domiciliarios ovonic-unisolar

    Energy Technology Data Exchange (ETDEWEB)

    Flores, J. Roberto; Agredano, Jaime; Munguia, Gonzalo; Lagunas, Javier; Huacuz, Jorge [Instituto de Investigaciones Electricas, Temixco, Morelos (Mexico); Brennan, Steve [Troy, MI (United States)

    2000-07-01

    In this work the first results of the evaluation and characterization of 30 solar home systems (SHS) Ovonic-Unisolar (16 of 30 W and 14 of 60 W of capacity) are presented. The components of the SHS are: One or two PV modules, a charge controller, a nickel metal-hybride (NiMH) battery with a nominal capacity of 85 Ah rated at 3 hours, two or four 8 W lamps, and a CD/CD converter for connecting a radio or a TV W/B a maximum power of 20W. Of all systems evaluated, 29 were installed in three communities of the Oaxaca State, and the other 2 are installed at the Instituto de Investigaciones Electricas (IIE) the laboratory as prototype systems. Before the systems installation at the rural communities, all of them were tested in laboratory. Eight systems installed in the field are being monitoring with data acquisition systems. The main motivation of this projects is to know the behavior of the NiMH battery in the SHS. [Spanish] En este trabajo se presentan los primeros resultados de la caracterizacion y evaluacion de 30 sistemas fotovoltaicos (FV) domiciliarios Ovonic-Unisolar (16 con capacidad de 30 W y 14 con capacidad de 60 W). Los sistemas estan integrados por uno o dos modulos FV de 30 W (dependiendo de la capacidad del sistema), un controlador de carga con termo-interuptor, una bateria del tipo niquel hidruros metalicos (NiHM) con capacidad de 85 Ah a una razon de descarga de 3 horas, 2 o 4 lamparas compactas de alta eficiencia de 8 W y un convertidor CD/CD que permite a los usuarios utilizar una radiograbadora y/o una television B/N con potencia no mayor a 20 W. De estos 30 sistemas FV, 28 se instalaron en 3 comunidades rurales del Estado de Oaxaca y los dos sistemas restantes se tienen instalados en el laboratorio FV del Instituto de Investigaciones Electricas (IIE) como sistemas testigos. Previo a la instalacion en campo, todos los sistemas fueron evaluados en el laboratorio para garantizar su operacion en las comunidades rurales. De los 28 sistemas instalados en

  8. 农产品电子商务供应链体系构建研究%Study on Construction of Farm Produce E-commerce Supply Chain System

    Institute of Scientific and Technical Information of China (English)

    刘丽华

    2012-01-01

    In this paper, we established an advanced and high-efficiency supply chain system and innovated the mode of the production and sales of agricultural products in a bid to satisfy the demand of the consumers for fresh green food.%电子商务时代,企业应立足农产品物流,应用电子商务,专注供应链管理,构建先进高效的供应链体系,创新农产品产销模式,以满足消费者对新鲜绿色食品的需求.

  9. An abstract approach to Loewner's chains

    CERN Document Server

    Arosio, Leandro; Hamada, Hidetaka; Kohr, Gabriela

    2010-01-01

    We present a new abstract construction of Loewner chains in one and several variables which holds also on hyperbolic complex manifolds and prove that there is essentially a one-to-one correspondence between evolution families of order $d$ and Loewner chains of the same order. As a consequence we obtain a Loewner-Kufarev generalized PDE on complex manifolds. We apply such results to the study of biholomorphic mappings from the unit ball of $\\C^n$.

  10. Construction management

    CERN Document Server

    Pellicer, Eugenio; Teixeira, José C; Moura, Helder P; Catalá, Joaquín

    2014-01-01

    The management of construction projects is a wide ranging and challenging discipline in an increasingly international industry, facing continual challenges and demands for improvements in safety, in quality and cost control, and in the avoidance of contractual disputes. Construction Management grew out of a Leonardo da Vinci project to develop a series of Common Learning Outcomes for European Managers in Construction. Financed by the European Union, the project aimed to develop a library of basic materials for developing construction management skills for use in a pan-European context. Focused exclusively on the management of the construction phase of a building project from the contractor's point of view, Construction Management covers the complete range of topics of which mastery is required by the construction management professional for the effective delivery of new construction projects. With the continued internationalisation of the construction industry, Construction Management will be required rea...

  11. Development of a biotinylated broad-specificity single-chain variable fragment antibody and a sensitive immunoassay for detection of organophosphorus pesticides.

    Science.gov (United States)

    Zhao, Fengchun; Tian, Yuan; Wang, Huimin; Liu, Jiye; Han, Xiao; Yang, Zhengyou

    2016-09-01

    Organophosphorus pesticides (OPs) are the most widely used pesticides in agriculture, and OP residues have been broadly reported in food and environmental samples. The aim of this study is to develop a recombinant antibody-based broad-specificity immunoassay for OPs. A phage display library was prepared from a mouse pre-immunized with a generic immunogen of OPs, and a single-chain variable fragment (scFv) antibody was selected. The selected scFv antibody was fused with biotin acceptor domain (BAD) and overexpressed as an inclusion body in Escherichia coli BL21 (DE3). Then, the protein was refolded by stepwise urea gradient dialysis and biotinylated in vitro by E. coli biotin ligase (BirA). Subsequently, the scFv-BAD protein was purified from the biotinylated system with high yield (66.7 mg L(-1)) and confirmed by SDS-PAGE and Western blot. Based on the biotinylated scFv-BAD, a sensitive and broad-specificity competitive indirect enzyme-linked immunosorbent assay (ciELISA) for detection of OPs was developed. The cross-reactivity (CR) studies demonstrated that the ciELISA described here exhibited the broadest detection spectrum for OPs up to now, and 30 OPs could be determined with 50 % inhibition value (IC50) values ranging from 19.4 to 515.2 ng mL(-1). Moreover, the developed ciELISA was used for the recovery study of the spiked samples and showed satisfactory recoveries. Graphical Abstract Schematic diagram of the development of biotinylated broad-specificity single-chain variable fragment antibody-based immunoassay for organophosphorus pesticides. PMID:27411546

  12. Spin chain sigma models with fermions

    Energy Technology Data Exchange (ETDEWEB)

    Hernandez, Rafael [Institut de Physique, Universite de Neuchatel, Breguet 1, CH-2000 Neuchatel (Switzerland)]. E-mail: rafael.hernandez@cern.ch; Lopez, [Departamento de Fisica Teorica C-XI and Instituto de Fisica Teorica C-XVI, Universidad Autonoma de Madrid, Cantoblanco, 28049 Madrid (Spain)

    2004-11-01

    The complete one-loop planar dilatation operator of N=4 supersymmetric Yang-Mills is isomorphic to the hamiltonian of an integrable PSU(2,2 vertical bar 4) quantum spin chain. We construct the non-linear sigma models describing the continuum limit of the SU(1|3) and SU(2|3) sectors of the complete N=4 chain. We explicitly identify the spin chain sigma model with the one for a superstring moving in AdS{sub 5} x S{sup 5} with large angular momentum along the five-sphere. (author)

  13. Decay Chain Deduction of Uranium Fission Products.

    Science.gov (United States)

    Guo, Huiping; Tian, Chenyang; Wang, Xiaotian; Lv, Ning; Ma, Meng; Wei, Yingguang

    2016-07-01

    Delayed gamma spectrum is the fingerprint of uranium materials in arms control verification technology. The decay chain is simplified into basic state linear chain and excitation state linear chain to calculate and analyze the delayed gamma spectra of fission products. Formulas of the changing rule for nuclide number before and after zero-time are deduced. The C program for calculating the delayed gamma ray spectra data is constructed, and related experiments are conducted to verify this theory. Through analysis of the delayed gamma counts of several nuclides, the calculated results are found to be consistent with experimental values. PMID:27218290

  14. 武汉软件服务外包产业价值链构建与价值提升策略%Industry Value Chain Construction and Value Promotion Strategy for Wuhan Software and Service Outsourcing

    Institute of Scientific and Technical Information of China (English)

    王梅源; 郑双怡; 熊兰兰

    2015-01-01

    产业价值链同时包含产业链和价值增值两个因素,有助于寻找与探索产业价值提升路径和策略。从软件生命周期、软件产品与服务、企业业务特征3个视角构建了软件服务外包产业价值链,针对武汉市产业发展现状和存在的问题,提出现阶段武汉软件服务外包产业应走一条“以技术升级和能力升级为主,创新研发与全球运营并重,产品升级与服务升级并重”的价值提升路径,并从区位优势、产业转移、在岸外包、龙头企业、创新发展5个方面提出产业价值提升的对策和建议。%Industry value chain includes two factors of the industrial chain and the value added at the same time, helps us find and explore the path and strategy of industry value upgrade. We build the software and service outsourcing industry value chain from three perspectives of the software life cycle, software products and services and enterprise business char‐acteristics. In view of the present situation and the existed question of Wuhan industry development, put forward a sug‐gestion that at present stage Wuhan software and service outsourcing industry must go a path of upgrade industries, that is "to upgrade technology and capability is given priority to, innovation development and global operations, product upgra‐ding and service upgrading pay equal attention to". We put forward the corresponding countermeasures and suggestions for industry value enhancement from five aspects of location advantage, industrial transfer, onshore outsourcing, leading en‐terprise, innovation and development.

  15. 湖北省农副产品冷链物流网络构建研究%Construction of Farm and Sideline Products Cold Chain Logistics Network in Hubei

    Institute of Scientific and Technical Information of China (English)

    旷玉玲; 海峰; 傅俊; 水璐; 方威

    2012-01-01

    将冷链物流网络从纵向上划分为冷链物流需求网络、冷链物流功能网络和冷链物流目标客户网络三个子网络.根据湖北省农副产品的发展现状,结合湖北省“两圈一带”的战略思路,提出了湖北省构建农副产品冷链物流网络的具体步骤、方法和建议.%This paper divides the cold chain logistics network vertically into three sub-systems, namely, demand network, function network and objective customer network. In view of the development of the farm and sideline products industry in Hubei and the strategic guideline of "two circles and one belt", it proposes some specfic procedures, measures and suggestions.

  16. COFFEE COMMODITY CHAIN

    OpenAIRE

    Tine Olsen; Brett Inder

    2008-01-01

    To explain the value added along the coffee commodity chain we propose and estimate a theoretical model of the coffee commodity chain. The theoretical model consists of four markets and five agents in the coffee commodity chain and predicts that prices in the coffee commodity chain move together but are also influenced by income, technology and production. A vector error correction model is used to test the theoretical predictions. In addition to the theoretical conclusions the empirical mode...

  17. The Global Value Chain

    DEFF Research Database (Denmark)

    Sørensen, Olav Jull

    The conference paper aims to develop the global value chain concept by including corporate internal value adding activities and competition to the basic framework in order to turn the global value chain into a strategic management tool......The conference paper aims to develop the global value chain concept by including corporate internal value adding activities and competition to the basic framework in order to turn the global value chain into a strategic management tool...

  18. Chain-Chain Based Routing Protocol

    Directory of Open Access Journals (Sweden)

    Samia A Ali

    2011-05-01

    Full Text Available Wireless sensor network (WSN is an emerging technology for monitoring physical world. WSNs consist of large numbers of sensor nodes operated by battery mostly in harsh environment. Thus energy conservation is a primary issue for organization of these sensor nodes. Another crucial issue is the data delivery time by sensor nodes to the sink node, especially in Military, medical fields, and security monitoring systems where minimum delay is desirable. Number of protocols has been proposed in the literature for routing. One of such protocols is the cluster based routing protocol LEACH (low energy adaptive clustering hierarchy. LEACH protocol organizes WSN into a set of clusters and a periodic voting for cluster head is performed in order to be evenly distributed among all the sensors of the WSN. This periodical cluster head voting in LEACH, however, consumes an amount of non-negligible energy and other resources. For energy conservation, PEGASIS (power efficient gathering in sensor information systems a near optimal chain-based protocol has been proposed, however, it is faced with the challenge of long delay for the transmitted data. Another routing protocol called CCM (Chain-Cluster based Mixed routing, which is mainly a hybrid of LEACH and PEGASIS is proposed, the consumed energy increases as network size increases. In this paper, we propose an efficient routing protocol called CCBRP (Chain-Chain based routing protocol, it achieves both minimum energy consumption and minimum delay. The CCBRP protocol mainly divides a WSN into a number of chains (Greedy algorithm is used to form each chain as in PEGSIS protocol and runs in two phases. In the first phase, sensor nodes in each chain transmit data to their chain leader nodes in parallel. In the second phase, all chain leader nodes form a chain (also, using Greedy algorithm and choose randomly a leader node then all chain leader nodes send their data to this chosen leader node. This chosen leader node

  19. Optimized Lentiviral Transduction Protocols by Use of a Poloxamer Enhancer, Spinoculation, and scFv-Antibody Fusions to VSV-G.

    Science.gov (United States)

    Anastasov, Nataša; Höfig, Ines; Mall, Sabine; Krackhardt, Angela M; Thirion, Christian

    2016-01-01

    Lentiviral vectors (LV) are widely used to successfully transduce cells for research and clinical applications. This optimized LV infection protocol includes a nontoxic poloxamer-based adjuvant combined with antibody-retargeted lentiviral particles. The novel poloxamer P338 demonstrates superior characteristics for enhancing lentiviral transduction over the best-in-class polybrene-assisted transduction. Poloxamer P338 exhibited dual benefits of low toxicity and high efficiency of lentiviral gene delivery into a range of different primary cell cultures. One of the major advantages of P338 is its availability in pharma grade and applicability as cell culture medium additive in clinical protocols. Lentiviral vectors pseudotyped with the vesicular stomatitis virus glycoprotein (VSV-G) can be produced to high titers and mediate high transduction efficiencies in vitro. For clinical applications the need for optimized transduction protocols, especially for transduction of primary T and stem cells, is high. The successful use of retronectin, the second lentivirus enhancer available as GMP material, requires the application of specific coating protocols not applicable in all processes, and results in the need of a relatively high multiplicity of infection (MOI) to achieve effective transduction efficiencies for hematopoietic cells (e.g., CD34+ hematopoietic stem cells). Cell specificity of lentiviral vectors was successfully increased by displaying different ratios of scFv-fused VSV-G glycoproteins on the viral envelope. The system has been validated with human CD30+ lymphoma cells, resulting in preferential gene delivery to CD30+ cells, which was increased fourfold in mixed cell cultures, by presenting scFv antibody fragments binding to respective surface markers. A combination of spinoculation and poloxamer-based chemical adjuvant increases the transduction of primary T-cells by greater than twofold. The combination of poloxamer-based and scFv-retargeted LVs increased

  20. Discrete Quantum Markov Chains

    CERN Document Server

    Faigle, Ulrich

    2010-01-01

    A framework for finite-dimensional quantum Markov chains on Hilbert spaces is introduced. Quantum Markov chains generalize both classical Markov chains with possibly hidden states and existing models of quantum walks on finite graphs. Quantum Markov chains are based on Markov operations that may be applied to quantum systems and include quantum measurements, for example. It is proved that quantum Markov chains are asymptotically stationary and hence possess ergodic and entropic properties. With a quantum Markov chain one may associate a quantum Markov process, which is a stochastic process in the classical sense. Generalized Markov chains allow a representation with respect to a generalized Markov source model with definite (but possibly hidden) states relative to which observables give rise to classical stochastic processes. It is demonstrated that this model allows for observables to violate Bell's inequality.

  1. Gushing metal chain

    Science.gov (United States)

    Belyaev, Alexander; Sukhanov, Alexander; Tsvetkov, Alexander

    2016-03-01

    This article addresses the problem in which a chain falls from a glass from some height. This phenomenon demonstrates a paradoxical rise of the chain over the glass. To explain this effect, an initial hypothesis and an appropriate theory are proposed for calculating the steady fall parameters of the chain. For this purpose, the modified Cayley's problem of falling chain given its rise due to the centrifugal force of upward inertia is solved. Results show that the lift caused by an increase in linear density at the part of chain where it is being bent (the upper part) is due to the convergence of the chain balls to one another. The experiments confirm the obtained estimates of the lifting chain.

  2. Usability Constructs

    DEFF Research Database (Denmark)

    Hertzum, Morten; Clemmesen, Torkil; Hornbæk, Kasper Anders Søren;

    2007-01-01

    Whereas research on usability predominantly employs universal definitions of the aspects that comprise usability, people experience their use of information systems through personal constructs. Based on 48 repertory-grid interviews, this study investigates how such personal constructs are affecte...

  3. scFv-based “grababody” as a general strategy to improve recruitment of immune effector cells to antibody-targeted tumors

    OpenAIRE

    Cai, Zheng; Fu, Ting; Nagai, Yasuhiro; Lam, Lian; Yee, Marla; Zhu, Zhiqiang; Zhang, Hongtao

    2013-01-01

    Recruitment of immune cells to tumor cells targeted by a therapeutic antibody can heighten the antitumor efficacy of the antibody. For example, p185her2/neu-targeting antibodies not only downregulate the p185her2/neu kinase (ERBB2) but also trigger complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) through the antibody Fc region. Here we describe a generalized strategy to improve immune cell recruitment to targeted cancer cells, using a modified scFv a...

  4. Autoconsumo de Energía Solar Fotovoltaica con Gestión Activa de la Demanda: El Sistema GeDELOS-FV

    OpenAIRE

    Caamaño Martín, Estefanía; Castillo Cagigal, Manuel; Masa Bote, Daniel; Gutiérrez Martín, Álvaro; Matallanas de Avila, Eduardo; Monasterio-Huelin Maciá, Felix; Jiménez Leube, Francisco Javier

    2010-01-01

    El sistema GeDELOS-FV es un ejemplo de valor añadido de la energía solar fotovoltaica que combina tecnología fotovoltaica híbrida de última generación con estrategias de Gestión de la Demanda Eléctrica. En este trabajo se ha configurado el sistema con el objetivo de maximizar la cantidad e energía fotovoltaica generada localmente y consumida por las cargas locales (autoconsumo), tanto de manera directa como indirecta mediante el uso de almacenamiento eléctrico. Se han realizado simulacion...

  5. Autoconsumo de energía solar fotovoltáica con gestión activa de la demanda:El sistema GEDELOS-FV

    OpenAIRE

    Caamaño Martín, Estefanía; Castillo Cagigal, Manuel; Masa Bote, Daniel; Gutiérrez Martín, Álvaro; Matallanas de Avila, Eduardo; Monasterio-Huelin Maciá, Felix; Jiménez Leube, Francisco Javier

    2010-01-01

    El sistema GeDELOS-FV es un ejemplo de valor añadido de la energía solar fotovoltaica que combina tecnología fotovoltaica híbrida de última generación con estrategias de Gestión de la Demanda Eléctrica. En este trabajo se ha configurado el sistema con el objetivo de maximizar la cantidad e energía fotovoltaica generada localmente y consumida por las cargas locales (autoconsumo), tanto de manera directa como indirecta mediante el uso de almacenamiento eléctrico. Se han realizado simulacione...

  6. CRITICAL EVENTS IN CONSTRUCTION PROCESS

    DEFF Research Database (Denmark)

    Jørgensen, Kirsten; Rasmussen, Grane Mikael Gregaard

    2009-01-01

    building process and includes all participants in the construction project. A general result from the analysis was that critical events that occurred when the site was not using Lean Construction evolved much longer than critical events that occurred in the period when Lean Construction was used. Another......Function failures, defects and poor communication are major problems in the construction industry. These failures and defects are caused by a row of critical events in the construction process. The purpose of this paper is to define “critical events” in the construction process and to investigate...... cause-effects of failures and defects in the construction industry by using an analytical approach (The bowtie model) which is developed in the accident research. Using this model clarifies the relationships within the chain of failures that causes critical events with undesirable consequences. In this...

  7. Self-assembly of linear [Mn II 2 Mn III ] units with end-on azido bridges: the construction of a ferromagnetic chain using S T = 7 high-spin trimers

    KAUST Repository

    Jiang, Yuan

    2015-01-01

    © The Royal Society of Chemistry 2015. The controlled organization of high-spin complexes into 1D coordination polymers is a challenge in molecular magnetism. In this work, we report a ferromagnetic Mn trimer Mn3(HL)2(CH3OH)6(Br)4·Br·(CH3OH)21 (H2L = 2-[(9H-fluoren-9-yl)amino]propane-1,3-diol) with the ground spin state of ST = 7 that can be assembled into a one-dimensional coordination chain [Mn3(HL)2(CH3OH)2(Br)4(N3)(H2O)·CH3OH]2 using azido bridging ligands. Interestingly, the ferromagnetic nature of 1 is well retained in 2. However, due to the negligible magnetic anisotropy in 1, both 1 and 2 do not show slow-relaxation of magnetization, which indicates that during the process of molecular assembly not only the intratrimer magnetic interaction but also the magnetic anisotropy of the trimer can be reserved.

  8. Side Chain Engineering in Solution-Processable Conjugated Polymers

    KAUST Repository

    Mei, Jianguo

    2014-01-14

    Side chains in conjugated polymers have been primarily utilized as solubilizing groups. However, these side chains have roles that are far beyond. We advocate using side chain engineering to tune a polymer\\'s physical properties, including absorption, emission, energy level, molecular packing, and charge transport. To date, numerous flexible substituents suitable for constructing side chains have been reported. In this Perspective article, we advocate that the side chain engineering approach can advance better designs for next-generation conjugated polymers. © 2013 American Chemical Society.

  9. Crater chains on Mercury

    Science.gov (United States)

    Shevchenko, V.; Skobeleva, T.

    After discovery of disruption comet Shoemaker-Levy 9 into fragment train before it's collision with Jupiter there was proposed that linear crater chains on the large satellites of Jupiter and on the Moon are impact scars of past tidally disrupted comets.It's known that radar images have revealed the possible presence of water ice deposits in polar regions of Mercury. Impacts by a few large comets seem to provide the best explanation for both the amount and cleanliness of the ice deposits on Mercury because they have a larger volatile content that others external sources, for example, asteroid. A number of crater chains on the surface of Mercury are most likely the impact tracks of "fragment trains" of comets tidally disrupted by Sun or by Mercury and are not secondary craters. Mariner 10 image set (the three Mariner 10 flybys in 1974-1975) was used to recognize the crater chains these did not associate with secondary crater ejecta from observed impact structures. As example, it can be shown such crater chain located near crater Imhotep and crater Ibsen (The Kuiper Quadrangle of Mercury). Resolution of the Mariner 10 image is about 0.54 km/pixel. The crater chain is about 50 km long. It was found a similar crater chain inside large crater Sophocles (The Tolstoj Quadrangle of Mercury). The image resolution is about 1.46 km/pixel. The chain about 50 km long is located in northen part of the crater. Image resolution limits possibility to examine the form of craters strongly. It seems the craters in chains have roughly flat floor and smooth form. Most chain craters are approximately circular. It was examined many images from the Mariner 10 set and there were identified a total 15 crater chains and were unable to link any of these directly to any specific large crater associated with ejecta deposits. Chain craters are remarkably aligned. All distinguished crater chains are superposed on preexisting formations. A total of 127 craters were identified in the 15 recognized

  10. Single-chain antibody-fragment M6P-1 possesses a mannose 6-phosphate monosaccharide-specific binding pocket that distinguishes N-glycan phosphorylation in a branch-specific manner†.

    Science.gov (United States)

    Blackler, Ryan J; Evans, Dylan W; Smith, David F; Cummings, Richard D; Brooks, Cory L; Braulke, Thomas; Liu, Xinyu; Evans, Stephen V; Müller-Loennies, Sven

    2016-02-01

    The acquisition of mannose 6-phosphate (Man6P) on N-linked glycans of lysosomal enzymes is a structural requirement for their transport from the Golgi apparatus to lysosomes mediated by the mannose 6-phosphate receptors, 300 kDa cation-independent mannose 6-phosphate receptor (MPR300) and 46 kDa cation-dependent mannose 6-phosphate receptor (MPR46). Here we report that the single-chain variable domain (scFv) M6P-1 is a unique antibody fragment with specificity for Man6P monosaccharide that, through an array-screening approach against a number of phosphorylated N-glycans, is shown to bind mono- and diphosphorylated Man6 and Man7 glycans that contain terminal αMan6P(1 → 2)αMan(1 → 3)αMan. In contrast to MPR300, scFv M6P-1 does not bind phosphodiesters, monophosphorylated Man8 or mono- or diphosphorylated Man9 structures. Single crystal X-ray diffraction analysis to 2.7 Å resolution of Fv M6P-1 in complex with Man6P reveals that specificity and affinity is achieved via multiple hydrogen bonds to the mannose ring and two salt bridges to the phosphate moiety. In common with both MPRs, loss of binding was observed for scFv M6P-1 at pH values below the second pKa of Man6P (pKa = 6.1). The structures of Fv M6P-1 and the MPRs suggest that the change of the ionization state of Man6P is the main driving force for the loss of binding at acidic lysosomal pH (e.g. lysosome pH ∼ 4.6), which provides justification for the evolution of a lysosomal enzyme transport pathway based on Man6P recognition. PMID:26503547

  11. Construction practice

    International Nuclear Information System (INIS)

    One has elaborated a virtual computer model (VRC) of construction of the AP1000 project reactor to demonstrate its viability, to improve the NPP project making it more easy for construction with simultaneous reduction of time, costs and risk of construction. The approach ensured time sequence of the 3-D visualization of NPP at the construction stages. The VRC ensures optimization of scheme implementation time period and specifies the basic costs. The VRC application offers essential advantages when planning construction of a nuclear power facility

  12. Targeting melanoma with immunoliposomes coupled to anti-MAGE A1 TCR-like single-chain antibody

    Directory of Open Access Journals (Sweden)

    Saeed M

    2016-03-01

    Full Text Available Mesha Saeed,1 Mandy van Brakel,2 Sara Zalba,1 Erik Schooten,2 Joost AP Rens,1 Gerben A Koning,1,† Reno Debets,2 Timo LM ten Hagen1 1Laboratory of Experimental Surgical Oncology, Section Surgical Oncology, Department of Surgery, Erasmus MC, Rotterdam, the Netherlands; 2Laboratory of Tumor Immunology, Department of Medical Oncology, Erasmus MC Cancer Institute, Rotterdam, the Netherlands †Dr Gerben A Koning passed away on December 29, 2015 Abstract: Therapy of melanoma using T-cells with genetically introduced T-cell receptors (TCRs directed against a tumor-selective cancer testis antigen (CTA NY-ESO1 demonstrated clear antitumor responses in patients without side effects. Here, we exploited the concept of TCR-mediated targeting through introduction of single-chain variable fragment (scFv antibodies that mimic TCRs in binding major histocompatibility complex-restricted CTA. We produced scFv antibodies directed against Melanoma AntiGEn A1 (MAGE A1 presented by human leukocyte antigen A1 (HLA-A1, in short M1/A1, and coupled these TCR-like antibodies to liposomes to achieve specific melanoma targeting. Two anti-M1/A1 antibodies with different ligand-binding affinities were derived from a phage-display library and reformatted into scFvs with an added cysteine at their carboxyl termini. Protein production conditions, ie, bacterial strain, temperature, time, and compartments, were optimized, and following production, scFv proteins were purified by immobilized metal ion affinity chromatography. Batches of pure scFvs were validated for specific binding to M1/A1-positive B-cells by flow cytometry. Coupling of scFvs to liposomes was conducted by employing different conditions, and an optimized procedure was achieved. In vitro experiments with immunoliposomes demonstrated binding of M1/A1-positive B-cells as well as M1/A1-positive melanoma cells and internalization by these cells using flow cytometry and confocal microscopy. Notably, the scFv with

  13. Targeting melanoma with immunoliposomes coupled to anti-MAGE A1 TCR-like single-chain antibody.

    Science.gov (United States)

    Saeed, Mesha; van Brakel, Mandy; Zalba, Sara; Schooten, Erik; Rens, Joost Ap; Koning, Gerben A; Debets, Reno; Ten Hagen, Timo Lm

    2016-01-01

    Therapy of melanoma using T-cells with genetically introduced T-cell receptors (TCRs) directed against a tumor-selective cancer testis antigen (CTA) NY-ESO1 demonstrated clear antitumor responses in patients without side effects. Here, we exploited the concept of TCR-mediated targeting through introduction of single-chain variable fragment (scFv) antibodies that mimic TCRs in binding major histocompatibility complex-restricted CTA. We produced scFv antibodies directed against Melanoma AntiGEn A1 (MAGE A1) presented by human leukocyte antigen A1 (HLA-A1), in short M1/A1, and coupled these TCR-like antibodies to liposomes to achieve specific melanoma targeting. Two anti-M1/A1 antibodies with different ligand-binding affinities were derived from a phage-display library and reformatted into scFvs with an added cysteine at their carboxyl termini. Protein production conditions, ie, bacterial strain, temperature, time, and compartments, were optimized, and following production, scFv proteins were purified by immobilized metal ion affinity chromatography. Batches of pure scFvs were validated for specific binding to M1/A1-positive B-cells by flow cytometry. Coupling of scFvs to liposomes was conducted by employing different conditions, and an optimized procedure was achieved. In vitro experiments with immunoliposomes demonstrated binding of M1/A1-positive B-cells as well as M1/A1-positive melanoma cells and internalization by these cells using flow cytometry and confocal microscopy. Notably, the scFv with nonenhanced affinity of M1/A1, but not the one with enhanced affinity, was exclusively bound to and internalized by melanoma tumor cells expressing M1/A1. Taken together, antigen-mediated targeting of tumor cells as well as promoting internalization of nanoparticles by these tumor cells is mediated by TCR-like scFv and can contribute to melanoma-specific targeting. PMID:27022262

  14. Construction supervision in construction project

    OpenAIRE

    Struna, Ervin

    2012-01-01

    Every construction project is a unique, complex process. Every completed structure is permanent act into space, and has therefore substantial influence upon social and natural environment. As a consequence, it is essential to provide professional supervision of the construction process that will ensure conformity of the construction with the project documentation and the building permit. The purpose of the thesis is to present the role of the construction supervision and the work of the s...

  15. Symmetry and resonance in periodic FPU chains

    NARCIS (Netherlands)

    Rink, B.

    2001-01-01

    The symmetry and resonance properties of the Fermi Pasta Ulam chain with periodic boundary conditions are exploited to construct a nearidentity transformation bring ing this Hamiltonian system into a particularly simple form This BirkhoGustavson normal form retains the symmetries of the original sy

  16. Preparation and functional studies of hydroxyethyl chitosan nanoparticles loaded with anti-human death receptor 5 single-chain antibody

    Directory of Open Access Journals (Sweden)

    Yang J

    2014-05-01

    Full Text Available Jingjing Yang,1,3,* Xiaoping Huang,1,3,* Fanghong Luo,1 Xiaofeng Cheng,3 Lianna Cheng,3 Bin Liu,4 Lihong Chen,2 Ruyi Hu,1,3 Chunyan Shi,1,3 Guohong Zhuang,1,3 Ping Yin2 1Anti-Cancer Research Center, Medical College, Xiamen University, Fujian, People's Republic of China, 2The Department of Pathology, Zhongshan Hospital, Xiamen University, Xiamen, People's Republic of China, 3Organ transplantation institution, Xiamen University, Xiamen, People's Republic of China, 4Jilin Vocational College of Industry and Technology, Jilin, People's Republic of China  *These authors contributed equally to this work Objective: To prepare hydroxyethyl chitosan nanoparticles loaded with anti-human death receptor 5 single-chain antibody, and study their characteristics, functions, and mechanisms of action. Materials and methods: The anti-human death receptor 5 single-chain antibody was constructed and expressed. Protein-loaded hydroxyethyl chitosan nanoparticles were prepared, and their size, morphology, particle-size distribution and surface zeta potential were measured by scanning electron microscopy and laser particle-size analysis. Mouse H22 hepatocellular carcinoma cells were cultured, and growth inhibition was examined using the CellTiter-Blue cell-viability assay. Flow cytometry and Hoechst 33342 were employed to measure cell apoptosis. Kunming mice with H22 tumor models were treated with protein-loaded hydroxyethyl chitosan nanoparticles, and their body weight and tumor size were measured, while hematoxylin and eosin staining was used to detect antitumor effects in vivo and side effects from tumors. Results: The protein-loaded hydroxyethyl chitosan nanoparticles had good stability; the zeta potential was -24.2±0.205, and the dispersion index was 0.203. The inhibition of the protein-loaded hydroxyethyl chitosan nanoparticles on H22 growth was both time- and dose-dependent. Increased expressions of active caspase 8, active caspase 3, and BAX were detected

  17. Supply Chain Management og Supply Chain costing

    DEFF Research Database (Denmark)

    Nielsen, Steen; Mortensen, Ole

    2002-01-01

    Formålet med denne artikel er at belyse de muligheder som ligger i at integrere virksomhedens økonomiske styring med begrebet Supply Chain Management (SCM). Dette søges belyst ved først at beskrive den teoretiske ramme, hvori SCM indgår. Herefter analyseres begrebet Supply Chain Costing (SCC) som...... Århus. Et resultat er, at via begrebet Supply Chain Costing skabes der mulighed for at måle logistikkædens aktiviteter i kr./øre. Anvendelsen af denne information har også strategisk betydning for at kunne vælge kunde og leverandør. Ved hjælp af integrationen skabes der også helt nye mulighed for at...

  18. Food supply chains

    OpenAIRE

    Zhou, Qian

    2011-01-01

    The dissertation analyses food waste in global supply chain. From the related managerial literature, the process of supply chain operation, from agriculture, manufacturing, warehouses, retailers to customers are explained clearly. Then the reasons and characteristics of food wastes in any point of food supply chain are analyzed. From some case studies and questionnaire investigation, some corresponding methods to reduce food waste are put forward in the following. Lastly, in terms of method s...

  19. Supply chain components

    Directory of Open Access Journals (Sweden)

    Vieraşu, T.

    2011-01-01

    Full Text Available In this article I will go through three main logistics components, which are represented by: transportation, inventory and facilities, and the three secondary logistical components: information, production location, price and how they determine performance of any supply chain. I will discuss then how these components are used in the design, planning and operation of a supply chain. I will also talk about some obstacles a supply chain manager may encounter.

  20. Supply Chain Collaboration

    OpenAIRE

    Meca, Ana; Timmer, Judith

    2008-01-01

    In this chaper, we have reviewed and surveyed the literature on supply chain collaboration. As mentioned above, the game theory models that include cooperative behaviour among retailers seem to be a natural framework to model cooperation (collaboration) in supply chains that consist of a supplier and a finite number of retailers. Various researchers in this area have already adopted several cooperative models dealing with supply chain coordination, and it is expected to see many more in the n...

  1. Supply chain components

    OpenAIRE

    Vieraşu, T.; Bălăşescu, M.

    2011-01-01

    In this article I will go through three main logistics components, which are represented by: transportation, inventory and facilities, and the three secondary logistical components: information, production location, price and how they determine performance of any supply chain. I will discuss then how these components are used in the design, planning and operation of a supply chain. I will also talk about some obstacles a supply chain manager may encounter.

  2. Sustainable Supply Chain Management

    OpenAIRE

    Geentjens, Marlies

    2013-01-01

    This project investigates the phenomena Sustainable Supply Chain Management. The point of departure is Supply Chain Management, which we analyse in order to lock into an understanding of the development that occur when implementing sustainability. In the quest for higher understanding of the interlocking mechanisms of supply chain management, we combine a theoretical approach to the developmnent of sustainability with some relevant case-examples (mainly depicting IKEA). This focal point g...

  3. Economy, market and chain

    OpenAIRE

    Sukkel, W.; Hommes, M.

    2009-01-01

    In their pursuit of growth and professionalisation, the Dutch organic sector focuses primarily on market development. But how do you stimulate the market for organic foods? This is the subject of many research projects concerning market, consumer preferences and the supply chain. These projects focus specifically at consumer purchasing behaviour, product development, supply chain formation and minimising cost price. As a rule, this research takes place in close cooperation with chain actors

  4. Structural Basis of Neutralization of the Major Toxic Component from the Scorpion Centruroides noxius Hoffmann by a Human-derived Single-chain Antibody Fragment

    Energy Technology Data Exchange (ETDEWEB)

    Canul-Tec, Juan Carlos; Riaño-Umbarila, Lidia; Rudiño-Piñera, Enrique; Becerril, Baltazar; Possani, Lourival D.; Torres-Larios, Alfredo (U. NAM)

    2011-08-09

    It has previously been reported that several single-chain antibody fragments of human origin (scFv) neutralize the effects of two different scorpion venoms through interactions with the primary toxins of Centruroides noxius Hoffmann (Cn2) and Centruroides suffusus suffusus (Css2). Here we present the crystal structure of the complex formed between one scFv (9004G) and the Cn2 toxin, determined in two crystal forms at 2.5 and 1.9 {angstrom} resolution. A 15-residue span of the toxin is recognized by the antibody through a cleft formed by residues from five of the complementarity-determining regions of the scFv. Analysis of the interface of the complex reveals three features. First, the epitope of toxin Cn2 overlaps with essential residues for the binding of {beta}-toxins to its Na+ channel receptor site. Second, the putative recognition of Css2 involves mainly residues that are present in both Cn2 and Css2 toxins. Finally, the effect on the increase of affinity of previously reported key residues during the maturation process of different scFvs can be inferred from the structure. Taken together, these results provide the structural basis that explain the mechanism of the 9004G neutralizing activity and give insight into the process of directed evolution that gave rise to this family of neutralizing scFvs.

  5. Value Chain Engineering

    DEFF Research Database (Denmark)

    Wæhrens, Brian Vejrum; Slepniov, Dmitrij

    2015-01-01

    This workbook is recommended for the attention of students of and managers in Danish small and medium sized enterprises (SMEs). Danish SMEs are currently facing a number of key challenges related to their position in global value chains. This book provides an insight into value chain management...... that may help these SMEs to occupy and sustain a competitive position in the value chain. It addresses this objective by introducing and discussing: • The concept of global value chains and its founding principles • The buyer-supplier relationships • Various SMEs operations configurations • Ideas for...

  6. Towards a synergy between project and supply chain management

    Directory of Open Access Journals (Sweden)

    Sanjay Sharma

    2012-10-01

    Full Text Available Construction projects are conventionally analyzed considering factors such as cost and completion time. The present paper focuses on the projects particularly dealing with the house or building. The paper aims at an objective of introducing a synergy between project and supply chain management. After a brief relevant literature review, major construction activities have been discussed and the associated synergy between project management and supply chain concepts has been explained. Applications of the supply chain concepts such as forward and reverse logistics along with the customer order decoupling point in the context of construction management are discussed as a result of the proposed development. For this purpose, a categorization of construction product is developed on the basis of land and building characteristics and it is the major contribution of the present paper. This is also expected to be useful for estimating lead time and availability of the product in the context of relevant supply chain, and an associated managerial implication.

  7. Model of service-oriented catering supply chain performance evaluation

    OpenAIRE

    Juanqiong Gou; Guguan Shen; Rui Chai

    2013-01-01

    Purpose: The aim of this paper is constructing a performance evaluation model for service-oriented catering supply chain. Design/methodology/approach: With the research on the current situation of catering industry, this paper summarized the characters of the catering supply chain, and then presents the service-oriented catering supply chain model based on the platform of logistics and information. At last, the fuzzy AHP method is used to evaluate the performance of service-oriented catering ...

  8. Self-assembly of Gold Nanoparticle Chains between Nanoelectrodes

    Institute of Scientific and Technical Information of China (English)

    Ning GU; Jian Hui LIAO; Lan HUANG; Zhi Rui GUO; Wei YU; Li Na XU; Jing WANG

    2006-01-01

    A novel strategy has been developed for construction of nanoparticle chains between nanoelectrodes with bifunctional molecules by taking advantage of linear aggregation of colloidal particles in organic solvents. As confirmed by scanning electron microscopy (SEM), an individual nanoparticle chain bridged the electrode pair. The present approach makes this technique to be cheap and may be applicable in microelectronic industry.

  9. An assay for the detection of grapevine leafroll-associated virus 3 using a single-chain fragment variable antibody.

    Science.gov (United States)

    Cogotzi, Laura; Giampetruzzi, Annalisa; Nölke, Greta; Orecchia, Martin; Elicio, Vito; Castellano, Maria Antonietta; Martelli, Giovanni P; Fischer, Rainer; Schillberg, Stefan; Saldarelli, Pasquale

    2009-01-01

    Grapevine leafroll-associated virus 3 (GLRaV-3) is a major pathogen of grapevine. A previously described single-chain fragment variable (scFv) antibody (scFvLR3), directed against the coat protein (CP) of GLRaV-3, was expressed in Escherichia coli and used to develop a diagnostic ELISA kit. The antibody was fused to the light chain constant domain of human immunoglobulin to create the bivalent reagent C(L)-LR3, which was purified from the periplasmic fraction, giving a yield of ~5 mg/l. The sensitivity of the reagent against recombinant GLRaV-3 CP was 0.1 ng. The sensitivity, specificity and durability of the reagent was similar to a commercial kit. The C(L)-LR3 showed a weak cross-reaction in immune electron microscopy assays to GLRaV-1 and -7, but not with the phylogenetically more distant GLRaV-2. A fully recombinant kit was developed with the inclusion of a recombinant GLRaV-3 CP expressed in bacteria, thus avoiding problems associated with virus propagation and purification. This system represents a rapid, simple, sensitive and standardized diagnostic protocol for GLRaV-3 detection. PMID:19082687

  10. Independent sets in chain cacti

    CERN Document Server

    Sedlar, Jelena

    2011-01-01

    In this paper chain cacti are considered. First, for two specific classes of chain cacti (orto-chains and meta-chains of cycles with h vertices) the recurrence relation for independence polynomial is derived. That recurrence relation is then used in deriving explicit expressions for independence number and number of maximum independent sets for such chains. Also, the recurrence relation for total number of independent sets for such graphs is derived. Finaly, the proof is provided that orto-chains and meta-chains are the only extremal chain cacti with respect to total number of independent sets (orto-chains minimal and meta-chains maximal).

  11. RFID Applied to Supply Chain Logistics in Disaster Recovery

    OpenAIRE

    Wang, Heng; Hubbard, Brian J; Hubbard, Sarah

    2014-01-01

    The purpose of this paper is to review recent developments to use RFID (Radio Frequency Identification) for supply chain logistics in the construction industry and identify potential applications of this technology for disaster recovery in construction operations. Natural disasters, terrorist attacks, and extreme weather conditions can cause significant damage to local communities and disruptions to the local supply chain. The effective movement of resources during disaster recovery operation...

  12. A Novel Evaluation Indicator System and Evaluation Method for Supply Chain Performance of Food Production

    OpenAIRE

    Zhai Xiyao; Ye Hankun

    2015-01-01

    Supply chain performance evaluation is a research hotspot and lies in the core status in supply chain management. The study presents a new evaluation indicator system and evaluation algorithm for supply chain performance. First, the balanced score card is used to construct an evaluation indicator system for supply chain performance evaluation through analyzing the basic principle and connotation characteristics of supply chain management; Second analytic hierarchy process and fuzzy comprehens...

  13. Sustainable Supply Chain Design

    DEFF Research Database (Denmark)

    Bals, Lydia; Tate, Wendy

    design for TBL sustainability. These are located in catastrophe-ridden Haiti and supported by the Germany-based investor company Yunus Social Business. Three supply chain archetypes combining physical and support chains are presented that focus on TBL sustainable outputs and outcomes. For SSCM research...

  14. Chain Reaction Polymerization.

    Science.gov (United States)

    McGrath, James E.

    1981-01-01

    The salient features and importance of chain-reaction polymerization are discussed, including such topics as the thermodynamics of polymerization, free-radical polymerization kinetics, radical polymerization processes, copolymers, and free-radical chain, anionic, cationic, coordination, and ring-opening polymerizations. (JN)

  15. Torus Construction

    OpenAIRE

    Valluri, Monica; Merritt, David

    1999-01-01

    The maximally compact representation of a regular orbit is in terms of its action-angle variables. Computing the map between a trajectory's Cartesian coordinates and its action-angle variables is called torus construction. This article reviews various approaches to torus construction and their application to galactic dynamics.

  16. Construction Management.

    Science.gov (United States)

    Abbott, James F.

    This article provides a detailed discussion of a team approach to building that involves a construction manager, an architect, and a contractor. Bidding methods are outlined; the major components in construction management -- value engineering and fast track scheduling -- and the use of performance specifications are discussed. The construction…

  17. Superstring construction

    CERN Document Server

    1989-01-01

    The book includes a selection of papers on the construction of superstring theories, mainly written during the years 1984-1987. It covers ten-dimensional supersymmetric and non-supersymmetric strings, four-dimensional heterotic strings and four-dimensional type-II strings. An introduction to more recent developments in conformal field theory in relation to string construction is provided.

  18. Integrated supply chain risk management

    OpenAIRE

    Riaan Bredell; Jackie Walters

    2007-01-01

    Integrated supply chain risk management (ISCRM) has become indispensable to the theory and practice of supply chain management. The economic and political realities of the modern world require not only a different approach to supply chain management, but also bold steps to secure supply chain performance and sustainable wealth creation. Integrated supply chain risk management provides supply chain organisations with a level of insight into their supply chains yet to be achieved. If correctly ...

  19. Supply Chain Connectivity: Enhancing Participation in the Global Supply Chain

    OpenAIRE

    Patalinghug, Epictetus E.

    2015-01-01

    Supply chain connectivity is vital for the efficient flow of trade among APEC economies. This paper reviews the literature and supply chain management, describes the barriers to enhancing participation in global supply chain, analyzes the various measures of supply chain performance, and suggests steps for the Philippines to fully reap the benefits of the global value chain.

  20. Asymptotic properties of quantum Markov chains

    International Nuclear Information System (INIS)

    The asymptotic dynamics of discrete quantum Markov chains generated by the most general physically relevant quantum operations is investigated. It is shown that it is confined to an attractor space in which the resulting quantum Markov chain is diagonalizable. A construction procedure of a basis of this attractor space and its associated dual basis of 1-forms is presented. It is applicable whenever a strictly positive quantum state exists which is contracted or left invariant by the generating quantum operation. Moreover, algebraic relations between the attractor space and Kraus operators involved in the definition of a quantum Markov chain are derived. This construction is not only expected to offer significant computational advantages in cases in which the dimension of the Hilbert space is large and the dimension of the attractor space is small, but it also sheds new light onto the relation between the asymptotic dynamics of discrete quantum Markov chains and fixed points of their generating quantum operations. Finally, we show that without any restriction our construction applies to all initial states whose support belongs to the so-called recurrent subspace. (paper)

  1. Targeting the active site of the placental isozyme of alkaline phosphatase by phage-displayed scFv antibodies selected by a specific uncompetitive inhibitor

    Directory of Open Access Journals (Sweden)

    Kala Mrinalini

    2005-12-01

    Full Text Available Abstract Background The isozymes of alkaline phosphatase, the tissue non-specific, intestinal and placental, have similar properties and a high degree of identity. The placental isozyme (PLAP is an oncofetal antigen expressed in several malignancies including choriocarcinoma, seminoma and ovarian carcinoma. We had earlier attempted to isolate PLAP-specific scFv from a synthetic human immunoglobulin library but were unable to do so, presumably because of the similarity between the isozymes. In this work, we have employed a PLAP-specific uncompetitive inhibitor, L-Phe-Gly-Gly, to select isozyme specific scFvs. An uncompetitive inhibitor binds to the enzyme in the presence of substrate and stabilizes the enzyme-substrate complex. Several uncompetitive inhibitors have varying degrees of isozyme specificity for human alkaline phosphatase isozymes. A specific uncompetitive inhibitor would be able to unmask conformational differences between the otherwise very similar molecules. Also, such inhibitors would be directed to regions at/close to the active site of the enzyme. In this work, the library was first incubated with PLAP and the bound clones then eluted by incubation with L-Phe-Gly-Gly along with the substrate, para-nitro phenyl phosphate (pNPP. The scFvs were then studied with regard to the biochemical modulation of their binding, isozyme specificity and effect on enzyme activity. Results Of 13 clones studied initially, the binding of 9 was inhibited by L-Phe-Gly-Gly (with pNPP and 2 clones were inhibited by pNPP alone. Two clones had absolute and 2 clones had partial specificity to PLAP. Two clones were cross-reactive with only one other isozyme. Three scFv clones, having an accessible His6-tag, were purified and studied for their modulation of enzyme activity. All the three scFvs inhibited PLAP activity with the kinetics of competitive inhibition. Cell ELISA could demonstrate binding of the specific scFvs to the cell surface expressed PLAP

  2. Construction management

    International Nuclear Information System (INIS)

    PEFP, Proton Engineering Frontier Project, puts its aim to building 100MeV 20mA linear proton accelerator which is national facility for NT, BT, IT, and future technologies, expected to boost up the national industry competitiveness. This R and D, Construction Management is in charge of the supportive works as site selection, architecture and engineering of conventional facilities, and overall construction management. The major goals of this work are as follows: At first, architecture and engineering of conventional facilities. Second, construction management, audit and inspection on construction of conventional facilities. Lastly, cooperation with the project host organization for adjusting technical issues of overall construction. In this research, We made a conceptual design, basic design and detail design of conventional facilities. Also, we made the engineering support for host site construction as follows : selection of project host organization and host site selection, construction technical work for project host organization and procedure management and impact analysis on the delay of project site selection

  3. Securitization of energy supply chains in China

    International Nuclear Information System (INIS)

    Highlights: • Three sources of energy security risks, namely sovereignty, robustness and resilience, affect China’s energy chains. • Energy security issues in China both have shaped and at the same time were shaped by ideas and institutions. • China remains rigid with equating ‘security’ with ‘national security’ and the notion of “national” is socially constructed. • Powerful actors, such as Chinese NOCs, inclined to interpret the problem so that it fits their preferred solution. • Securitization of any energy supply chains results from their historical roots, system properties and institutional agents. - Abstract: Energy policies in China, the world’s largest energy consumer, are an important factor in shaping the global energy system. While scholars agree that energy security is a major driver of China’s energy policies, there is insufficient understanding of what exactly constitutes China’s energy security from the policy perspective. We apply recent insights from the Global Energy Assessment, particularly the idea of vital energy systems, and the securitization theory to propose a framework for explaining China’s energy security policies in their historic evolution. We pay specific attention to explaining how particular energy supply chains are constructed and securitized. We draw data from over 300 Chinese and over 100 English publications and 30 interviews with energy officials and experts in China. We demonstrate that China’s focus on vulnerabilities of its oil supply chain at the expense of improving the reliability of domestic electricity supply is not accidental. It has its roots in historic events, properties of energy systems, as well as the presence of powerful institutional agents interested in securitizing the oil supply chain but not other vital energy systems. We suggest that this focus on the oil supply chain is likely to be maintained in the future, possibly accompanied by increasing concerns over natural gas

  4. Understanding the supply chain

    Directory of Open Access Journals (Sweden)

    Aćimović Slobodan

    2006-01-01

    Full Text Available Supply chain management represents new business philosophy and includes strategically positioned and much wider scope of activity in comparison with its "older brother" - management of logistics. Philosophy of the concept of supply chain is directed to more coordination of key business functions of every link in distribution chain in the process of organization of the flow of both goods and information, while logistic managing instruments are focused on internal optimum of flows of goods and information within one company. Applying the concept of integrated supply chain among several companies makes the importance of operative logistics activity even greater on the level of one company, thus advancing processes of optimum and coordination within and between different companies and confirms the importance of logistics performances for the company’s profitability. Besides the fact that the borders between companies are being deleted, this concept of supply chain in one distribution channel influences increasing of importance of functional, i.e. traditional business managing approaches but instead it points out the importance of process managing approaches. Although the author is aware that "there is nothing harder, more dangerous and with uncertain success, but to find a way for introducing some novelties (Machiavelli, it would be even his additional stimulation for trying to bring closer the concept and goals of supply chain implementation that are identified in key, relevant, modern, theoretical and consulting approaches in order to achieve better understanding of the subject and faster implementation of the concept of supply chain management by domestic companies.

  5. SUMO chains: polymeric signals.

    Science.gov (United States)

    Vertegaal, Alfred C O

    2010-02-01

    Ubiquitin and ubiquitin-like proteins are conjugated to a wide variety of target proteins that play roles in all biological processes. Target proteins are conjugated to ubiquitin monomers or to ubiquitin polymers that form via all seven internal lysine residues of ubiquitin. The fate of these target proteins is controlled in a chain architecture-dependent manner. SUMO (small ubiquitin-related modifier) shares the ability of ubiquitin to form chains via internal SUMOylation sites. Interestingly, a SUMO-binding site in Ubc9 is important for SUMO chain synthesis. Similar to ubiquitin-polymer cleavage by USPs (ubiquitin-specific proteases), SUMO chain formation is reversible. SUMO polymers are cleaved by the SUMO proteases SENP6 [SUMO/sentrin/SMT3 (suppressor of mif two 3)-specific peptidase 6], SENP7 and Ulp2 (ubiquitin-like protease 2). SUMO chain-binding proteins including ZIP1, SLX5/8 (synthetic lethal of unknown function 5/8), RNF4 (RING finger protein 4) and CENP-E (centromere-associated protein E) have been identified that interact non-covalently with SUMO chains, thereby regulating target proteins that are conjugated to SUMO multimers. SUMO chains play roles in replication, in the turnover of SUMO targets by the proteasome and during mitosis and meiosis. Thus signalling via polymers is an exciting feature of the SUMO family. PMID:20074033

  6. Construction mathematics

    CERN Document Server

    Virdi, Surinder

    2006-01-01

    Taking a starting point below that of GCSE level, by assuming no prior mathematical knowledge, Surinder Virdi and Roy Baker take the reader step by step through the mathematical requirements for Level 2 and 3 Building and Construction courses.Unlike the majority of basic level maths texts available, this book focuses exclusively on mathematics as it is applied in actual construction practice. As such, topics specific to the construction industry are presented, as well as essential areas for Level 2 craft NVQs - for example, costing calculations, labor costs, cost of materials and setting out o

  7. Enablers of Innovation in the Construction Material Industry

    DEFF Research Database (Denmark)

    Wandahl, Søren; Lassen, Astrid Heidemann; Jacobsen, Alexia;

    2014-01-01

    The construction material industry is often acknowledged as slightly more innovative than the overall construction industry and could hence serve as a valuable learning place for how innovation could flourish in the construction industry. Construction is viewed as network or supply chain based...

  8. Project Decision Chain

    DEFF Research Database (Denmark)

    Rolstadås, Asbjørn; Pinto, Jeffrey K.; Falster, Peter;

    2015-01-01

    To add value to project performance and help obtain project success, a new framework for decision making in projects is defined. It introduces the project decision chain inspired by the supply chain thinking in the manufacturing sector and uses three types of decisions: authorization, selection......, and plan decision. A primitive decision element is defined where all the three decision types can be accommodated. Each task in the primitive element can in itself contain subtasks that in turn will comprise new primitive elements. The primitive elements are nested together in a project decision chain....

  9. Quantum Markov Chain Mixing and Dissipative Engineering

    DEFF Research Database (Denmark)

    Kastoryano, Michael James

    2012-01-01

    This thesis is the fruit of investigations on the extension of ideas of Markov chain mixing to the quantum setting, and its application to problems of dissipative engineering. A Markov chain describes a statistical process where the probability of future events depends only on the state of the sy....... Finally, we consider three independent tasks of dissipative engineering: dissipatively preparing a maximally entangled state of two atoms trapped in an optical cavity, dissipative preparation of graph states, and dissipative quantum computing construction.......This thesis is the fruit of investigations on the extension of ideas of Markov chain mixing to the quantum setting, and its application to problems of dissipative engineering. A Markov chain describes a statistical process where the probability of future events depends only on the state...... (stationary states). The aim of Markov chain mixing is to obtain (upper and/or lower) bounds on the number of steps it takes for the Markov chain to reach a stationary state. The natural quantum extensions of these notions are density matrices and quantum channels. We set out to develop a general mathematical...

  10. Worldwide construction

    International Nuclear Information System (INIS)

    The paper lists major construction projects in worldwide processing and pipelining, showing capacities, contractors, estimated costs, and time of construction. The lists are divided into refineries, petrochemical plants, sulfur recovery units, gas processing plants, pipelines, and related fuel facilities. This last classification includes cogeneration plants, coal liquefaction and gasification plants, biomass power plants, geothermal power plants, integrated coal gasification combined-cycle power plants, and a coal briquetting plant

  11. Nested Bethe ansatz for "all" closed spin chains

    OpenAIRE

    Belliard, S.; Ragoucy, E.

    2008-01-01

    We present in an unified and detailed way the Nested Bethe Ansatz for closed spin chains based on Y(gl(n)), Y(gl(m|n)), U_q(gl(n)) or U_q(gl(m|n)) (super)algebras, with arbitrary representations (i.e. `spins') on each site of the chain. In particular, the case of indecomposable representations of superalgebras is studied. The construction extends and unifies the results already obtained for spin chains based on Y(gl(n)) or U_q(gl(n)) and for some particular super-spin chains. We give the Beth...

  12. Supply chain risk management

    Directory of Open Access Journals (Sweden)

    Christian Hollstein

    2013-03-01

    Full Text Available Background: Supply chain risk management increasingly gains prominence in many international industries. In order to strengthen supply chain structures, processes, and networks, adequate potentials for risk management need to be built (focus on effective logistics and to be utilized (focus on efficient logistics. Natural-based disasters, such as the case of Fukushima, illustrate how crucial risk management is. Method: By aligning a theoretical-conceptual framework with empirical-inductive findings, it may be hypothesized that logistical systems do have a positive effect on supply chain risk management activities.  Result/conclusion:  Flexibility and capacity, as well as redundancy and standardization, are often viewed as being conflictionary. It shows, however, that in the light of supply chain risk management, those factors may yield a common benefit if proper logistics systems are applied.  

  13. Editorial: Supply Chain Management

    Directory of Open Access Journals (Sweden)

    Aidonis, D.

    2012-01-01

    Full Text Available This special issue has followed up the 2nd Olympus International Conference on Supply Chains held on October 5-6, 2012, in Katerini, Greece. The Conference was organized by the Department of Logistics of Alexander Technological Educational Institution (ATEI of Thessaloniki, in collaboration with the Laboratory of Quantitative Analysis, Logistics and Supply Chain Management of the Department of Mechanical Engineering, Aristotle University of Thessaloniki (AUTH. During the 2-Days Conference more than 50 research papers were presented covering the following thematic areas: (i Business Logistics, (ii Transportation, Telematics and Distribution Networks, (iii Green Logistics, (iv Information and Communication Technologies in Supply Chain Management, and (v Services and Quality. Three keynote invited speakers addressed interesting issues for the Humanitarian Logistics, Green Supply Chains of the Agrifood Sector and the Opportunities and Prospects of Greek Ports chaired Round Tables with other Greek and Foreign Scientists and Specialists.

  14. Environmental Retail Supply Chains

    DEFF Research Database (Denmark)

    Kotzab, Herbert; Munch, Hilde; de Faultrier, Birgitte;

    2011-01-01

    Purpose – The purpose of this paper is to develop a scale that evaluates the environmental elements in retail supply chains and to examine the environmental supply chain management initiatives of the world's largest 100 retailing companies. Design/methodology/approach – The empirical evaluation has...... which were grouped into eight categories; they refer to “fundamental environmental attitude”, “use of energy”, “use of input material”, “product”, “packaging”, “transport”, “consumption” and “waste”. The level of environmental supply chain management can be characterised as very operational and very...... short-term oriented (green operations). Long-term oriented green design initiatives were hardly observed. Furthermore, the specific environmental activities of three retailers from Denmark, France and the UK were compared. Research limitations/implications – The empirical study investigates supply chain...

  15. ACCELERATING CONSTRUCTION INDUSTRY DEVELOPMENT

    Directory of Open Access Journals (Sweden)

    Mohan M. Kumaraswamy

    2006-06-01

    Full Text Available The needs for construction industry development are initially viewed from the broader perspectives of imperatives for infrastructure development and national development. All these are clearly more critical in developing countries. A non-exhaustive set of potential drivers and common barriers to construction industry development is identified from previous research. These suggest the usefulness of consolidating a cluster of recent proposals and exercises aiming at (a construction organization development in terms of an over-arching management support system model, as well as improved information and knowledge management; and (b project team development in the context of relationally integrated teams and supply chains, joint risk management and ‘technology and knowledge exchange’ in joint ventures, as well as longer term public private partnerships. These apparently disparate research thrusts are threaded together into a pattern that may inspire, if not feed, specific research and development (R & D agendas for construction industry development in different countries according to their own priorities, constraints and stages of infrastructure and national development.

  16. Supply Chain Collaboration

    OpenAIRE

    Guy Rodrigue Ghomsi 1982

    2015-01-01

    Companies are now progressively looking beyond themselves to come up with ways to improve revenue and profit margins. This has resulted in a heightened focus on the efficiency of the supply chains. Inefficient supply chains are being perceived as cavities that can deprive the firm from its cash flows when inventory is not required and sales revenue when product is not available at the point of sales. This has stimulated development of business models that intend to leverage and enhance the su...

  17. Variance bounding Markov chains

    OpenAIRE

    Roberts, Gareth O.; Jeffrey S. Rosenthal

    2008-01-01

    We introduce a new property of Markov chains, called variance bounding. We prove that, for reversible chains at least, variance bounding is weaker than, but closely related to, geometric ergodicity. Furthermore, variance bounding is equivalent to the existence of usual central limit theorems for all L2 functionals. Also, variance bounding (unlike geometric ergodicity) is preserved under the Peskun order. We close with some applications to Metropolis–Hastings algorithms.

  18. Supply chain risk management

    OpenAIRE

    Christian Hollstein; Frank Himpel

    2013-01-01

    Background: Supply chain risk management increasingly gains prominence in many international industries. In order to strengthen supply chain structures, processes, and networks, adequate potentials for risk management need to be built (focus on effective logistics) and to be utilized (focus on efficient logistics). Natural-based disasters, such as the case of Fukushima, illustrate how crucial risk management is. Method: By aligning a theoretical-conceptual framework with empirical-induct...

  19. Editorial: Supply Chain Management

    OpenAIRE

    Aidonis, D.; Folinas, D.; Triantafillou, D.

    2012-01-01

    This special issue has followed up the 2nd Olympus International Conference on Supply Chains held on October 5-6, 2012, in Katerini, Greece. The Conference was organized by the Department of Logistics of Alexander Technological Educational Institution (ATEI) of Thessaloniki, in collaboration with the Laboratory of Quantitative Analysis, Logistics and Supply Chain Management of the Department of Mechanical Engineering, Aristotle University of Thessaloniki (AUTH). During the 2-Days Conference m...

  20. Construction and Characterization of a Naive Camelus Bactrianus Variable Domain of Heavy Chain of Heavy-Chain Antibody(VHH)Yeast Two-Hybrid Library%双峰驼天然重链抗体可变区酵母双杂交文库的构建与鉴定

    Institute of Scientific and Technical Information of China (English)

    胡湘云; 高小龙; 付向晶; 刘丹丹; 范文涛; 王燕平; 杜恩岐; 王兴龙; 党如意

    2015-01-01

    构建双峰驼天然重链抗体可变区(VHH)酵母双杂交文库,并对文库质量进行鉴定.采集未经免疫的6月龄雌性双峰驼外周血、骨髓、脾和淋巴结,并从外周血中分离淋巴细胞.抽提总RNA,反转录为cDNA,用2对引物经过2轮PCR扩增得到400 bp左右的双峰驼VHH片段.根据Clontech公司Mate&PlateTM library construction system使用手册,将带有同源臂的VHH片段与线性化的pGADT7-Rec质粒共转化Y187酵母感受态细胞,转化产物涂布SD/-Leu平板,30℃倒置培养3~5 d后,收集平板上的所有克隆,即为双峰驼天然重链抗体可变区酵母双杂交文库.对文库的滴度、重组效率及多样性进行鉴定.结果显示,本研究构建的双峰驼天然重链抗体可变区酵母双杂交文库库容为2.07×107,滴度为7.6×108 cfu·mL-1.随机挑取47个独立克隆进行菌落PCR鉴定,43个含有VHH片段,重组率为91.5%;选取其中19个测序,均为独立克隆,且多样性好.该研究成功构建双峰驼天然重链抗体可变区酵母双杂交文库,且文库质量满足要求,为进一步获得特定抗原的VHH奠定了基础.

  1. Issues on Supply Chain Management

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Although supply chain is regarded as an integrated pr ocess, actually, researchers and practitioners have primarily investigated the v arious processes of the supply chain individually. The competitive field in most markets requires well-designed integrated supply chain instead of individual o perations. In this paper, some traditional thinking of supply chain is given fir st, then SCM (Supply Chain Management) is introduced, which views the entire sup ply chain as a whole rather than individual process a...

  2. The effect of internalizing human single chain antibody fragment on liposome targeting to epithelioid and sarcomatoid mesothelioma

    Science.gov (United States)

    Iyer, Arun K.; Su, Yang; Feng, Jinjin; Lan, Xiaoli; Zhu, Xiaodong; Liu, Yue; Gao, Dongwei; Seo, Youngho; VanBrocklin, Henry F.; Broaddus, V. Courtney; Liu, Bin; He, Jiang

    2011-01-01

    Immunoliposomes (ILs) anchored with internalizing human antibodies capable of targeting all subtypes of mesothelioma can be useful for targeted imaging and therapy of this malignant disease. The objectives of this study were to evaluate both the in vitro and in vivo tumor targeted internalization of novel internalizing human single chain antibody (scFv) anchored ILs on both epithelioid (M28) and sarcomatoid (VAMT-1) subtypes of human mesothelioma. ILs were prepared by post-insertion of mesothelioma-targeting human scFv (M1) onto preformed liposomes and radiolabeled with 111In (111In-IL-M1), along with control non-targeted liposomes (111In-CL). Incubation of 111In-IL-M1 with M28, VAMT-1, and a control non-tumorigenic cell-line (BPH-1) at 37°C for 24 h revealed efficient binding and rapid internalization of ILs into both subtypes of tumor cells but not into the BPH-1 cells; internalization accounted for approximately 81-94% of total cell accumulation in mesothelioma cells compared to 37-55% in control cells. In tumor bearing mice intravenous (i.v.) injection of 111In-IL-M1 led to remarkable tumor accumulation: 4 % and 4.7% injected dose per gram (% ID/g) for M28 and VAMT-1 tumors, respectively, 48 h after injection. Furthermore, tumor uptake of 111In-IL-M1 in live xenograft animal models was verified by single photon emission computed tomography (SPECT/CT). In contrast, i.v. injection of 111In-CL in tumor-bearing mice revealed very low uptake in both subtypes of mesothelioma, 48 h after injection. In conclusion, M1 scFv-anchored ILs showed selective tumor targeting and rapid internalization into both epithelioid and sarcomatoid subtypes of human mesothelioma, demonstrating its potential as a promising vector for enhanced tumor drug targeting. PMID:21255833

  3. Biodistribution and tumor imaging of an anti-CEA single-chain antibody-albumin fusion protein

    Energy Technology Data Exchange (ETDEWEB)

    Yazaki, Paul J. [Division of Cancer Immunotherapeutics and Tumor Immunology, Beckman Research Institute, City of Hope, Duarte, CA 91010 (United States)], E-mail: pyazaki@coh.org; Kassa, Thewodros; Cheung, Chia-wei; Crow, Desiree M. [Division of Cancer Immunotherapeutics and Tumor Immunology, Beckman Research Institute, City of Hope, Duarte, CA 91010 (United States); Sherman, Mark A. [Division of Information Sciences, Beckman Research Institute, City of Hope, Duarte, CA 91010 (United States); Bading, James R.; Anderson, Anne-Line J.; Colcher, David; Raubitschek, Andrew [Division of Cancer Immunotherapeutics and Tumor Immunology, Beckman Research Institute, City of Hope, Duarte, CA 91010 (United States)

    2008-02-15

    Albumin fusion proteins have demonstrated the ability to prolong the in vivo half-life of small therapeutic proteins/peptides in the circulation and thereby potentially increase their therapeutic efficacy. To evaluate if this format can be employed for antibody-based imaging, an anticarcinoembryonic antigen (CEA) single-chain antibody(scFv)-albumin fusion protein was designed, expressed and radiolabeled for biodistribution and imaging studies in athymic mice bearing human colorectal carcinoma LS-174T xenografts. The [{sup 125}I]-T84.66 fusion protein demonstrated rapid tumor uptake of 12.3% injected dose per gram (ID/g) at 4 h that reached a plateau of 22.7% ID/g by 18 h. This was a dramatic increase in tumor uptake compared to 4.9% ID/g for the scFv alone. The radiometal [{sup 111}In]-labeled version resulted in higher tumor uptake, 37.2% ID/g at 18 h, which persisted at the tumor site with tumor: blood ratios reaching 18:1 and with normal tissues showing limited uptake. Based on these favorable imaging properties, a pilot [{sup 64}Cu]-positron emission tomography imaging study was performed with promising results. The anti-CEA T84.66 scFv-albumin fusion protein demonstrates highly specific tumor uptake that is comparable to cognate recombinant antibody fragments. The radiometal-labeled version, which shows lower normal tissue accumulation than these recombinant antibodies, provides a promising and novel platform for antibody-based imaging agents.

  4. Construction safety

    CERN Document Server

    Li, Rita Yi Man

    2013-01-01

    A close-to-ideal blend of suburb and city, speedy construction of towers of Babylon, the sparkling proportion of glass and steel buildings’ facade at night showcase the wisdom of humans. They also witness the footsteps, sweats and tears of architects and engineers. Unfortunately, these signatures of human civilizations are swathed in towering figures of construction accidents. Fretting about these on sites, different countries adopt different measures on sites. This book firstly sketches the construction accidents on sites, followed by a review on safety measures in some of the developing countries such as Bermuda, Egypt, Kuwait and China; as well as developed countries, for example, the United States, France and Singapore. It also highlights the enormous compensation costs with the courts’ experiences in the United Kingdom and Hong Kong.

  5. Construction Physics

    OpenAIRE

    Bertelsen, Sven; Henrich, Guilherme; Koskela, Lauri; Rooke, John

    2007-01-01

    At the 14th annual conference in the International Group for Lean Construction in 2006 the authors presented the concept of construction as a process being fed by a number of flows out of which one is the critical one deciding the speed with which the process takes place. This contribution takes the idea further and sets out by discussing production in general and the underlying mental flow models associated with this understanding. It then suggests a new model for the understanding of the na...

  6. Handbook of Markov chain Monte Carlo

    CERN Document Server

    Brooks, Steve

    2011-01-01

    ""Handbook of Markov Chain Monte Carlo"" brings together the major advances that have occurred in recent years while incorporating enough introductory material for new users of MCMC. Along with thorough coverage of the theoretical foundations and algorithmic and computational methodology, this comprehensive handbook includes substantial realistic case studies from a variety of disciplines. These case studies demonstrate the application of MCMC methods and serve as a series of templates for the construction, implementation, and choice of MCMC methodology.

  7. Test automation for Markov Chain Usage Models

    OpenAIRE

    Bettinotti, Adriana M.; Garavaglia, Mauricio

    2011-01-01

    Statistical testing with Markov Chain Usage Models is an effective method to be used by programmers and testers during web sites development, to guarantee the software reliability. The JUMBL software works on this models; it supports model construction with the TML language and analysis, tests generation and execution and analysis of tests results. This paper is targeted at test automation for web sites development with JUMBL and JWebUnit.

  8. Phasic Triplet Markov Chains.

    Science.gov (United States)

    El Yazid Boudaren, Mohamed; Monfrini, Emmanuel; Pieczynski, Wojciech; Aïssani, Amar

    2014-11-01

    Hidden Markov chains have been shown to be inadequate for data modeling under some complex conditions. In this work, we address the problem of statistical modeling of phenomena involving two heterogeneous system states. Such phenomena may arise in biology or communications, among other fields. Namely, we consider that a sequence of meaningful words is to be searched within a whole observation that also contains arbitrary one-by-one symbols. Moreover, a word may be interrupted at some site to be carried on later. Applying plain hidden Markov chains to such data, while ignoring their specificity, yields unsatisfactory results. The Phasic triplet Markov chain, proposed in this paper, overcomes this difficulty by means of an auxiliary underlying process in accordance with the triplet Markov chains theory. Related Bayesian restoration techniques and parameters estimation procedures according to the new model are then described. Finally, to assess the performance of the proposed model against the conventional hidden Markov chain model, experiments are conducted on synthetic and real data. PMID:26353069

  9. Construction work

    CERN Multimedia

    2004-01-01

    Construction work on building 179 will start on the 16th February 2004 and continue until November 2004. The road between buildings 179 and 158 will temporarily become a one way street from Route Democrite towards building 7. The parking places between buildings 179 and 7 will become obsolete. The ISOLDE collaboration would like to apologize for any inconveniences.

  10. Supply chain quality management

    Directory of Open Access Journals (Sweden)

    Hannan Amoozad Mahdiraji

    2012-10-01

    Full Text Available In recent years, there are several methods introduced for the improvement of operational performances. Total quality management and supply chain management are two methods recommended for this purpose. These two approaches have been studied in most researches separately, while they have objectives in common, and this makes them a strategic means, which can be used, simultaneously. Total quality management and supply chain management play significant roles to increase the organizational competitiveness power. Moreover, they have only one purpose that is customer satisfaction, and they are different only on their approaches to reach their objectives. In this research, we aim to study both approaches of quality management and supply chain, their positive increasing effects that may be generated after their integration. For this purpose, the concept and definitions of each approach is studied, independently, their similarities and differences are recognized, and finally, the advantages of their integration are introduced.

  11. Essentials of supply chain management

    CERN Document Server

    Hugos, Michael H

    2011-01-01

    The latest thinking, strategies, developments, and technologies to stay current in supply chain management Presenting the core concepts and techniques of supply chain management in a clear, concise and easily readable style, the Third Edition of Essentials of Supply Chain Management outlines the most crucial tenets and concepts of supply chain management.

  12. Building the resilient supply chain.

    OpenAIRE

    Christopher, Martin; Peck, Helen

    2004-01-01

    In today’s uncertain and turbulent markets, supply chain vulnerability has become an issue of significance for many companies. As supply chains become more complex as a result of global sourcing and the continued trend to ‘leaning-down’, supply chain risk increases. The challenge to business today is to manage and mitigate that risk through creating more resilient supply chains.

  13. Supply-Chain Optimization Template

    Science.gov (United States)

    Quiett, William F.; Sealing, Scott L.

    2009-01-01

    The Supply-Chain Optimization Template (SCOT) is an instructional guide for identifying, evaluating, and optimizing (including re-engineering) aerospace- oriented supply chains. The SCOT was derived from the Supply Chain Council s Supply-Chain Operations Reference (SCC SCOR) Model, which is more generic and more oriented toward achieving a competitive advantage in business.

  14. Fuzzy Markov chains: uncertain probabilities

    OpenAIRE

    James J. Buckley; Eslami, Esfandiar

    2002-01-01

    We consider finite Markov chains where there are uncertainties in some of the transition probabilities. These uncertainties are modeled by fuzzy numbers. Using a restricted fuzzy matrix multiplication we investigate the properties of regular, and absorbing, fuzzy Markov chains and show that the basic properties of these classical Markov chains generalize to fuzzy Markov chains.

  15. Integrated supply chain risk management

    Directory of Open Access Journals (Sweden)

    Riaan Bredell

    2007-11-01

    Full Text Available Integrated supply chain risk management (ISCRM has become indispensable to the theory and practice of supply chain management. The economic and political realities of the modern world require not only a different approach to supply chain management, but also bold steps to secure supply chain performance and sustainable wealth creation. Integrated supply chain risk management provides supply chain organisations with a level of insight into their supply chains yet to be achieved. If correctly applied, this process may optimise management decision-making and assist in the protection and enhancement of shareholder value.

  16. Model of service-oriented catering supply chain performance evaluation

    Directory of Open Access Journals (Sweden)

    Juanqiong Gou

    2013-03-01

    Full Text Available Purpose: The aim of this paper is constructing a performance evaluation model for service-oriented catering supply chain. Design/methodology/approach: With the research on the current situation of catering industry, this paper summarized the characters of the catering supply chain, and then presents the service-oriented catering supply chain model based on the platform of logistics and information. At last, the fuzzy AHP method is used to evaluate the performance of service-oriented catering supply chain. Findings: With the analysis of the characteristics of catering supply chain, we construct the performance evaluation model in order to guarantee the food safety, logistics efficiency, price stability and so on. Practical implications: In order to evolve an efficient and effective service supply chain, it can not only used to own enterprise improvement, but also can be used for selecting different customers, to choose a different model of development. Originality/value: This paper has a new definition of service-oriented catering supply chain. And it offers a model to evaluate the performance of this catering supply chain.

  17. Modelling Constructs

    DEFF Research Database (Denmark)

    Kindler, Ekkart

    2009-01-01

    There are many different notations and formalisms for modelling business processes and workflows. These notations and formalisms have been introduced with different purposes and objectives. Later, influenced by other notations, comparisons with other tools, or by standardization efforts, these...... notations have been extended in order to increase expressiveness and to be more competitive. This resulted in an increasing number of notations and formalisms for modelling business processes and in an increase of the different modelling constructs provided by modelling notations, which makes it difficult...... to compare modelling notations and to make transformations between them. One of the reasons is that, in each notation, the new concepts are introduced in a different way by extending the already existing constructs. In this chapter, we go the opposite direction: We show that it is possible to add...

  18. Constructive Interactions

    OpenAIRE

    Lemoisson, Philippe; Cerri, Stefano A.; Mahe, Serge-André; Sallantin, Jean

    2003-01-01

    The new paradigm of "knowledge construction using experiential based and collaborative learning approaches" is an outstanding opportunity for interdisciplinary research. This document is an attempt to introduce and exemplify as much as possible using the lexicon of "social sciences", considerations and tools belonging to "artificial intelligence" (eg.:the machine learning tradition). In the paper we first draw a conceptual framework for rational agents in conversational interaction; then we u...

  19. Plastic value chains

    DEFF Research Database (Denmark)

    Baxter, John; Wahlstrom, Margareta; Zu Castell-Rüdenhausen, Malin;

    2014-01-01

    Optimizing plastic value chains is regarded as an important measure in order to increase recycling of plastics in an efficient way. This can also lead to improved awareness of the hazardous substances contained in plastic waste, and how to avoid that these substances are recycled. As an example...

  20. Innovation in Supply Chains

    DEFF Research Database (Denmark)

    Maier, Maximilian; Korbel, Jakob; Brem, Alexander

    2015-01-01

    . Moreover, along with the fourth industrial revolution – industry 4.0 – new technologies such as cyber physical systems are fast gaining popularity. Hence, based on the analysis of relevant literature, we further develop the supply chain committee model, developed by Kaluza et al. (2003) to demonstrate how...

  1. Heavy chain only antibodies

    DEFF Research Database (Denmark)

    Moghimi, Seyed Moein; Rahbarizadeh, Fatemeh; Ahmadvand, Davoud;

    2013-01-01

    Unlike conventional antibodies, heavy chain only antibodies derived from camel contain a single variable domain (VHH) and two constant domains (CH2 and CH3). Cloned and isolated VHHs possess unique properties that enable them to excel conventional therapeutic antibodies and their smaller antigen...

  2. METRODOS: Meteorological preprocessor chain

    DEFF Research Database (Denmark)

    Astrup, P.; Mikkelsen, T.; Deme, S.

    2001-01-01

    - heat flux related measurement, e.g. a temperature gradient, are used to give local values of friction velocity and Monin-Obukhov length plus an estimate of the mixing height. The METRODOS meteorological preprocessor chain is an integral part of the RODOS - Real Time On Line Decision Support - program...

  3. Supply chain reliability modelling

    Directory of Open Access Journals (Sweden)

    Eugen Zaitsev

    2012-03-01

    Full Text Available Background: Today it is virtually impossible to operate alone on the international level in the logistics business. This promotes the establishment and development of new integrated business entities - logistic operators. However, such cooperation within a supply chain creates also many problems related to the supply chain reliability as well as the optimization of the supplies planning. The aim of this paper was to develop and formulate the mathematical model and algorithms to find the optimum plan of supplies by using economic criterion and the model for the probability evaluating of non-failure operation of supply chain. Methods: The mathematical model and algorithms to find the optimum plan of supplies were developed and formulated by using economic criterion and the model for the probability evaluating of non-failure operation of supply chain. Results and conclusions: The problem of ensuring failure-free performance of goods supply channel analyzed in the paper is characteristic of distributed network systems that make active use of business process outsourcing technologies. The complex planning problem occurring in such systems that requires taking into account the consumer's requirements for failure-free performance in terms of supply volumes and correctness can be reduced to a relatively simple linear programming problem through logical analysis of the structures. The sequence of the operations, which should be taken into account during the process of the supply planning with the supplier's functional reliability, was presented.

  4. Shared Value Chain Design

    DEFF Research Database (Denmark)

    Bals, Lydia; Tate, Wendy L.

    In Sustainable Supply Chain Management (SSCM) research still the classic economic perspective is the dominating perspective, although the triple bottom line (including economic, social and ecological) is well accepted. The theoretical foundation for the paper is Stakeholder Theory. Case studies o...

  5. Exploration Supply Chain Simulation

    Science.gov (United States)

    2008-01-01

    The Exploration Supply Chain Simulation project was chartered by the NASA Exploration Systems Mission Directorate to develop a software tool, with proper data, to quantitatively analyze supply chains for future program planning. This tool is a discrete-event simulation that uses the basic supply chain concepts of planning, sourcing, making, delivering, and returning. This supply chain perspective is combined with other discrete or continuous simulation factors. Discrete resource events (such as launch or delivery reviews) are represented as organizational functional units. Continuous resources (such as civil service or contractor program functions) are defined as enabling functional units. Concepts of fixed and variable costs are included in the model to allow the discrete events to interact with cost calculations. The definition file is intrinsic to the model, but a blank start can be initiated at any time. The current definition file is an Orion Ares I crew launch vehicle. Parameters stretch from Kennedy Space Center across and into other program entities (Michaud Assembly Facility, Aliant Techsystems, Stennis Space Center, Johnson Space Center, etc.) though these will only gain detail as the file continues to evolve. The Orion Ares I file definition in the tool continues to evolve, and analysis from this tool is expected in 2008. This is the first application of such business-driven modeling to a NASA/government-- aerospace contractor endeavor.

  6. CHAINE DES ROTISSEUR @ HILTON

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    The annual gala dinner of Chaine des Rotisseurs was hosted by Hilton Beijing. The “Ballet Dinner” - a great creation for both art and fine dining lovers was taken place in varies places within the hotel, including hotel lobby, 3rd floor and lobby lounge.

  7. Construction and Biological Effects in vitro of Double-Directed Tissue-Specific HSV-tk/GCV Anti-Tumor System

    Institute of Scientific and Technical Information of China (English)

    WANGQian; ZHUHuifen; CAOLimin; YUBing; ZHOUHuarong; SHENGuanxin

    2003-01-01

    Objective: To construct a specifically targeted gene delivery and expression system, and to investigate the special killing effect of the HSV-tk/GCV system on human liver cancer cells in vitro.Methods: The anti-transferrin receptor (TfR) ScFv-GAL4 fusion protein expression vector ScFv-GAL4-pET28a and the eukaryotic expression plasmid pEBAF/tk-GAL4rec were constructed by recombinant DNA technology. After the induction by IPTG, we obtained the anti-TfR ScFv-GAL4 fusion protein as delivery vector to transfect pEBAF/tk-GAL4rec into the human liver cancer cell lines HepG2 and SMMC7721 and the human lung cancer cell line A549 that overexpress TfR via receptor-mediated endocytosis. The positive cell clones were selected by hygromycin and named HepG2/tk, SMMC7721/tk and A549/tk,respectively. Cell killing after GCV application was determined by MTT. Results: The correct structure of the ScFv-Gal4 fusion protein and the plasmid pEBAF/tk-GAL4rec was confirmed by double enzyme digestion, SDS-PAGE and sequencing. HepG2/tk cells that express alphafetoprotein (AFP) to high levels(845 ng/ml) were very sensitive to GCV, while SMMC7721/tk cells that express AFP at low levels (2ng/ml) and AFP-negative A549/tk cells were only slightly or not sensitive to GCV. Conclusion: The double-directed and tissue-specific HSV-tk/GCV anti-tumor system shows good targeting to tumor cells.

  8. Constructing Catalonia

    Directory of Open Access Journals (Sweden)

    Bill Phillips

    2009-01-01

    Full Text Available Catalonia, in common with other nations, has long b een concerned with the question of identity and difference. Its problemati c relationship with Spain has led to an emphasis on differentiating itself from its larger neighbour (if we are to accept, as most Spaniards do not, that Catalonia is not Spain, a s ituation complicated by the loss of the Spanish colonies of Cuba and The Philippines in 189 8, and the Spanish Civil War and subsequent dictatorship from 1936 to 1976. Beginnin g in the late nineteenth century, the construction of a Catalan identity followed a simil ar route to that taken by other European nations such as England, Ireland and, inde ed, Spain, including an emphasis on rural values, activities and the countryside, and t he conversion of specifically local traditions into national past times. It is only in the last ten years or so that this model of Catalan identity has been recognised for what it is – a model constructed and encouraged for and by specific nationalist politica l interests. Ironically, Catalonia’s identity abroad has also been constructed and manip ulated for political purposes, but from quite a different perspective. Orwell’s /Homag e to Catalonia/ (1938 narrates an extremely blinkered version of the Spanish Civil Wa r which has achieved iconic status as a result of cold war politics. Subsequent portra yals of the Spanish Civil War – Valentine Cunningham’s /The Penguin Book of Spanish Civil War Verse/ (ed., Penguin, 1980, or Ken Loach’s 1995 film /Land and F reedom/ base their arguments unquestioningly on /Homage to Catalonia/, perpetuat ing a view of the nation’s recent history that is both reductive and inaccurate

  9. Constructing Costs

    OpenAIRE

    Donner, Henrik af

    2000-01-01

    The economic analysis of building contracts is an unexplored field within Law and Economics. This paper makes an attempt to cast some light over the subject and encourage to more research within the field. The main purpose has been to describe why the actual price in a public construction project often turns out to be higher than the contract price and offer a way of handle that risk. In the paper a model is set up that shows an optimal contract given expectations on actual price and gaps in ...

  10. Impact of supply chain relational interactions on information sharing

    OpenAIRE

    Zouaghi, I.; A. Spalanzani; Gunasekaran, A.

    2011-01-01

    The environment complexity in which supply chain members are evolving leads them to adopt opposite strategies in a constructive way. In this sense companies tend to create value by cooperation and, at the same time, capture value by competition. This ago-antagonistic behavior generates a relational dynamics in which companies perform on a set of factors to improve their position within the chain. This enables them to progress towards common interests while ensuring on their own ones. In the p...

  11. Connectivity in Logistics and Supply Chain Management: A Framework

    OpenAIRE

    Hoffman, Karolin; Hellström, Daniel

    2008-01-01

    This paper seeks to investigate the connectivity construct in logistics and supply chain management literature. A review of existing literature is conducted regarding connectivity on a broad basis. The review is based on a general search in the ABI-Inform database for articles containing the twin search phrases of connectivity and logistics, and connectivity and supply chain management. An initial analysis of articles found resulted in a total of 195 articles, of which 76 are found to be rele...

  12. Interaction between supply chain risk and organizational performance

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    To offer a theoretic guidance to practice, on the basis of the work in an emerging strand of supply chain management and by matching the constructs of performance and risk together, a framework between supply chain risk management and organizational performance is developed. The framework includes a series of mutually associated processes as follows: risk sources and profiles, risk and performance drives, risk and performance consequences, risk management responses, and risk and performance outcomes. Furthe...

  13. On almost-periodic points of a topological Markov chain

    International Nuclear Information System (INIS)

    We prove that a transitive topological Markov chain has almost-periodic points of all D-periods. Moreover, every D-period is realized by continuously many distinct minimal sets. We give a simple constructive proof of the result which asserts that any transitive topological Markov chain has periodic points of almost all periods, and study the structure of the finite set of positive integers that are not periods.

  14. Markov Chains as Tools for Jazz Improvisation Analysis

    OpenAIRE

    Franz, David Matthew

    1998-01-01

    This thesis describes an exploratory application of a statistical analysis and modeling technique (Markov chains) for the modeling of jazz improvisation with the intended subobjective of providing increased insight into an improviser's style and creativity through the postulation of quantitative measures of style and creativity based on the constructed Markovian analysis techniques. Using Visual Basic programming language, Markov chains of orders one to three are created using transcriptio...

  15. Innovation Across the Supply Chain

    DEFF Research Database (Denmark)

    Druehl, Cheryl; Carrillo, Janice; Hsuan, Juliana

    Innovation is an integral part of every firm’s ongoing operations. Beyond product innovation, supply chain innovations offer a unique source of competitive advantage. We synthesize recent research on innovation in the supply chain, specifically, innovative supply chain processes such as collabora...... collaborations with customers and suppliers to create new product and process innovations, open innovation, and globally distributed innovation. We conclude with potential areas for future research.......Innovation is an integral part of every firm’s ongoing operations. Beyond product innovation, supply chain innovations offer a unique source of competitive advantage. We synthesize recent research on innovation in the supply chain, specifically, innovative supply chain processes such as...

  16. Schistosoma japonicum:construction of phage display antibody library and its application in the immunodiagnosis of infection

    Institute of Scientific and Technical Information of China (English)

    陈代雄; 何蔼; 詹希美; 俞慕华; 雷智刚; 孟锦绣; 李卓雅; 梁瑜; 张瑞琳

    2004-01-01

    Background A monoclonal antibody would be an effective tool for the detection of circulating antigens in the serum of patients with schistosomiasis, but the traditional way of producing monoclonal antibodies is not cost-effective. The objective of this study was to find a new method for the large-scale production of monoclonal antibodies against Schistosoma japonicum (Sj).Methods A phage display antibody library for Sj was constructed. To obtain a single-chain variable fragment antibody (scFv) against Sj, the library was screened with metabolic antigens from adult Sj worms (Sj-MAg) using enzyme-linked immunosorbent assay. The soluble scFvs selected were used to detect Sj antigens in the serum of acute and chronic schistosomiasis patients.Results Six positive clones with good reactivity to Sj-MAg were obtained from the phage display antibody library of about 1.07×106 individual clones. Only two of these six clones bound specifically to Sj-MAg and were chosen for further analysis. Specific soluble anti-Sj-MAg scFvs were produced by inducing the 2 clones with isopropyl-D-thiogalactopyranoside. The characteristics of the scFvs were then determined. The results of Western blot showed that these scFvs could bind to Sj-MAg specifically and had a molecular weight of about 31 kD. When testing serum from schistosomiasis patients with one of the two specific scFvs, its sensitivity was found to be 60% and 37% in acute and chronic patients, respectively, with a specificity of 90%. When the two specific scFvs were combined, their sensitivity was found to be 75% and 57% in acute and chronic patients, respectively, with a specificity of 85%.Conclusions The results indicate that the scFvs are potentially useful for the diagnosis of schistosomiasis. The library construction also provides a useful tool for the further screening of other antibodies for both diagnostic and immunotherapeutic applications and for epitope analysis and vaccine design.

  17. Decision-Making for Supply Chain Integration Supply Chain Integration

    CERN Document Server

    Lettice, Fiona; Durowoju, Olatunde

    2012-01-01

    Effective supply chain integration, and the tight co-ordination it creates, is an essential pre-requisite for successful supply chain management.  Decision-Making for Supply Chain Integration is a practical reference on recent research in the area of supply chain integration focusing on distributed decision-making problems. Recent applications of various decision-making tools for integrating supply chains are covered including chapters focusing on: •Supplier selection, pricing strategy and inventory decisions in multi-level supply chains, •RFID-enabled distributed decision-making, •Operational risk issues and time-critical decision-making for sensitive logistics nodes, Modelling end to end processes to improve supply chain integration, and •Integrated systems to improve service delivery and optimize resource use. Decision-Making for Supply Chain Integration provides an insight into the tools and methodologies of this field with support from real-life case studies demonstrating successful application ...

  18. Volumes of chain links

    CERN Document Server

    Kaiser, James; Rollins, Clint

    2011-01-01

    Agol has conjectured that minimally twisted n-chain links are the smallest volume hyperbolic manifolds with n cusps, for n at most 10. In his thesis, Venzke mentions that these cannot be smallest volume for n at least 11, but does not provide a proof. In this paper, we give a proof of Venzke's statement. The proof for n at least 60 is completely rigorous. The proof for n between 11 and 59 uses a computer calculation, and can be made rigorous for manifolds of small enough complexity, using methods of Moser and Milley. Finally, we prove that the n-chain link with 2m or 2m+1 half-twists cannot be the minimal volume hyperbolic manifold with n cusps, provided n is at least 60 or |m| is at least 8, and we give computational data indicating this remains true for smaller n and |m|.

  19. Demand chain management-integrating marketing and supply chain management

    OpenAIRE

    Jüttner, Uta; Christopher, Martin; Baker, Susan

    2007-01-01

    This paper endorses demand chain management as a new business model aimed at creating value in today's marketplace, and combining the strengths of marketing and supply chain competencies. Demand chain design is based on a thorough market understanding and has to be managed in such a way as to effectively meet differing customer needs. Based on a literature review as well as the findings from a co-development workshop and focus group discussions with marketing and supply chain professionals, a...

  20. Supply Chain Initiatives Database

    Energy Technology Data Exchange (ETDEWEB)

    None

    2012-11-01

    The Supply Chain Initiatives Database (SCID) presents innovative approaches to engaging industrial suppliers in efforts to save energy, increase productivity and improve environmental performance. This comprehensive and freely-accessible database was developed by the Institute for Industrial Productivity (IIP). IIP acknowledges Ecofys for their valuable contributions. The database contains case studies searchable according to the types of activities buyers are undertaking to motivate suppliers, target sector, organization leading the initiative, and program or partnership linkages.