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Volume 26 Issue 4
August 2011
    Citation: Hong Tian, Yan Cheng, Jin-yang Wu, Jian-hui He, You-jun Shang, Xiang-tao Liu. Application of GP5 Protein to Develop Monoclonal Antibody against Porcine Reproductive and Respiratory Syndrome Virus * .VIROLOGICA SINICA, 2011, 26(4) : 267-272.  http://dx.doi.org/10.1007/s12250-011-3198-5
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    Application of GP5 Protein to Develop Monoclonal Antibody against Porcine Reproductive and Respiratory Syndrome Virus *

    cstr: 32224.14.s12250-011-3198-5

    • State Key Laboratory of Veterinary Etiological Biology, National Foot and Mouth Disease Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China

    • Corresponding author: Xiang-tao Liu, hnxiangtao@hotmail.com
    • Received Date: 27 April 2011
      Accepted Date: 27 June 2011
      Available online: 01 August 2011

      Fund Project: Chinese National Technology Research and Development Program 863 ProgramChinese National Technology Research and Development Program 2006AA10A204

    • In this study, a panel of monoclonal antibodies (mAbs) against Porcine reproductive and respiratory syndrome virus(PRRSV), named as 8C9 and4B4, were produced by fusing SP2/0 myeloma cells and spleen cells of BALB/c mice immunized with the PRRSV (TCID50=5.5), screened by the indirect ELISA and subjected to several limiting dilutions. mAbs were then identified by biological characterization. Among the two fusion cell strains, 8C9 belonged to the IgG1 subclass and 4B4 belonged to the IgG2a subclass. The titers in cell culture supernatant and abdomen liquor reached to 1:104and 1:105, respectively. The specificity test indicated that the two cells had specific reactions for the PRRSV and GP5 protein respectively, and no reaction with Classical swine fever virus (CSFV) or Swine vesicular disease virus (SVDV). The molecular weights of the heavy chain and light chain were about 45.0 kDa and 25.0 kDa, respectively. In neutralization activity tests, the results showed that the prepared mAb 4B4 can protect 50% of cells with no CPE in dilution up to 1:512, but mAB 8C9 has no neutralization activities to PRRSV.
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      1. Chang C C, Yoon K J, Zimmerman J J, et al. 2002. Evolution of porcine reproductive and respiratory syndrome virus during sequential passages in pigs. J Virol, 76(10):4750-4763.
          doi: 10.1128/JVI.76.10.4750-4763.2002

      2. Chen J, Liu T, Zhu C G, et al. 2006. Genetic variation of Chinese PRRSV strains based on ORF5 sequence. Biochem Genet, 44, 425-435.

      3. Davis J M, Pennington J E, Kubler A M, et al. 1982. A simple, single-step technique for selecting and cloning hybridomas for the production of monoclonal antibodies. J Immunol Methods, 50, 161-171.
          doi: 10.1016/0022-1759(82)90222-8

      4. Gao Z Q, Guo X, Yang H C. 2004. Genomic characterization of two Chinese isolates of porcine respiratory and reproductive syndrome virus. Arch Virol, 149, 1341-1351.

      5. Golding S M, Hedger R S, Talbot P. 1976. Radial immunodiffusion and serum neutralization techniques for the assay of antibodies to swine vesicular disease. Res Vet Sci, 20:142-147.

      6. Hanada K, Suzuki Y, Nakane T, et al. 2005. The origin and evolution of porcine reproductive and respiratory syndrome viruses. Mol Biol Evol, 22(11):2131-2134.
          doi: 10.1093/molbev/msi208

      7. Murtaugh M P, Elam M R, Kakach L T. 1995. Comparison of the structural protein coding sequences of the VR-2332 and Lelystad virus strains of the PRRS virus. Arch Virol, 1451-1460.

      8. Polson D D, Marsh W E, Dial G D. 1992. Financial evaluation and decision making in the swine breeding herd. Vet Clin North Am Food Anim Pract, 8:725-747.
          doi: 10.1016/S0749-0720(15)30713-1

      9. Tong L, Jun D, Jun S, et al. 2009. Application of VP1 Protein to Develop Monoclonal Antibody against Foot-and-mouth Disease Virus Asia1 Type. Virol Sin, 24(3):215-220.
          doi: 10.1007/s12250-009-2986-z

      10. Wissink E H, van Wijk H A, Kroese M V, et al. 2003. The major envelope protein, GP5, of a European porcine reproductive and respiratory syndrome virus contains a neutralization epitope in its N-terminal ectodomain. J Gen Virol, 84: 1535-1543.
          doi: 10.1099/vir.0.18957-0

      11. Yan C, Hong T, Jian H, et al.2011. Indirect ELISA with recombinant GP5 for detecting antibodies to porcine reproductive and respiratory syndrome virus. Virol Sin, 26(1):61-66.
          doi: 10.1007/s12250-011-3154-9

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      Application of GP5 Protein to Develop Monoclonal Antibody against Porcine Reproductive and Respiratory Syndrome Virus *

        Corresponding author: Xiang-tao Liu, hnxiangtao@hotmail.com
      • State Key Laboratory of Veterinary Etiological Biology, National Foot and Mouth Disease Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China
      • Received Date: 27 April 2011
      • Accepted Date: 27 June 2011
      Fund Project:  Chinese National Technology Research and Development Program 863 ProgramChinese National Technology Research and Development Program 2006AA10A204

      Abstract: In this study, a panel of monoclonal antibodies (mAbs) against Porcine reproductive and respiratory syndrome virus(PRRSV), named as 8C9 and4B4, were produced by fusing SP2/0 myeloma cells and spleen cells of BALB/c mice immunized with the PRRSV (TCID50=5.5), screened by the indirect ELISA and subjected to several limiting dilutions. mAbs were then identified by biological characterization. Among the two fusion cell strains, 8C9 belonged to the IgG1 subclass and 4B4 belonged to the IgG2a subclass. The titers in cell culture supernatant and abdomen liquor reached to 1:104and 1:105, respectively. The specificity test indicated that the two cells had specific reactions for the PRRSV and GP5 protein respectively, and no reaction with Classical swine fever virus (CSFV) or Swine vesicular disease virus (SVDV). The molecular weights of the heavy chain and light chain were about 45.0 kDa and 25.0 kDa, respectively. In neutralization activity tests, the results showed that the prepared mAb 4B4 can protect 50% of cells with no CPE in dilution up to 1:512, but mAB 8C9 has no neutralization activities to PRRSV.

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