Development of monoclonal antibodies against nucleoprotein of Newcastle disease virus and establishment of a quantitative double-antibody sandwich ELISA for NDV antigen / 中国人兽共患病学报

ABSTRACT

We developed the monoclonal antibodies against nucleoprotein (NP) of Newcastle disease virus (NDV),and established a double antibody sandwich ELISA method for quantitative determination of NP antigen of NDV (NDV NP ELISA).The recombination NP protein derived from strain F48E9 of NDV were prepared and used to immunize BLAB/c mice.The mouse splenic cells from immunized mice were fused with SP2/0 cells to generate monoclonal antibodies (mAb).The NDV NP specific mAbs were paired to establish a double antibody sandwich ELISA method.The performance of the NDV NP ELISA was evaluated,including specificity,sensitivity,precision,accuracy and linearity.The correlation between the ELISA and PFU virus titer was analyzed by regression analysis method.Two monoclonal antibodies 3C10 and 4E7 were selected to establish double antibody sandwich ELISA for NP antigen of NDV.The linearity and performance of the NDV NP ELISA was characterized.The detection linearity fell in the range of 0.015-0.250 μg/mL (R2 =0.997 4).The detection limit of the assay was 0.015 μg/mL.The recovery was between 88.4％ and 106.01％;the variation coefficient was below 3.4％.In testing of 50 NDV virus samples,this assay performed well and correlated comparably with PFU virus titer (R2 =0.920 9).The NDV NP ELISA for quantitative detection of NDV is a reliable quantifiable assay for detection of NDV NP protein;it provides a new approach for rapid and quantitative detection of Newcastle disease virus.