Scientia Agricultura Sinica ›› 2012, Vol. 45 ›› Issue (7): 1399-1405.doi: 10.3864/j.issn.0578-1752.2012.07.019

• VETERINARY SCIENCE • Previous Articles     Next Articles

Production and Characterization of Anti-PrP Polyclonal Antibody by PRNP Gene Knockout Goat

 WANG  Xue-Bin, YU  Hui-Qing, WU  Xiao-Dong, XU  Xu-Jun, CHEN  Jian-Quan, CHENG  Guo-Xiang   

  1. 1.上海转基因研究中心，上海 201210
    2.上海杰隆生物工程股份有限公司，上海 201210
    3.中国动物卫生与流行病学中心, 青岛 266032，山东
  • Received:2011-11-22 Online:2012-04-01 Published:2012-02-06

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Abstract: 【Objective】The study was conducted in order to produce and characterize anti-PrP polyclonal antibodies by prion protein knockout (PRNP-/-) goat.【Method】Using polymerase chain reaction (PCR) method, the DNA sequence encoding the goat mature PrP (Prion Protein) was amplified from goat DNA. It was then cloned into the vector pET-22b to express recombinant PrP protein. After purification, the recombinant PrP was used as antigen to immunize PRNP-/- goat for producing anti-PrP antibodies. ELISA and Western blot were conducted to characterize the titer and specificity of polyclonal antibodies.【Result】A large scale of anti-PrP polyclonal antibodies was successfully produced by PRNP-/- goat. ELISA assay revealed that the titer of the polyclonal antibodies against PrP was as high as 1﹕25 600. Western blot test showed that the antibody was not only able to react with the native prion proteins (PrPC) of mouse, cattle and goat brain, but also could combine with misfolded prion proteins (PrPSc) from mouse brain.【Conclusion】Ablation of the prion protein(PrP) gene in goat facilitates the production of a large scale anti-PrP polyclonal antibodies, the produced antibody can be widely used in detecting PrPC and PrPSc derived from a variety of animals.

Key words: PrPC,  , PrPSc, polyclonal antibodies, PRNP-/- goat

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