Expression of NP protein of crimean-congo hemorrhagic fever virus in E.coli and preparation of monoclonal antibodies against NP protein
Chinese Journal of Health Laboratory Technology(2009)
摘要
Objective:To express NP protein of crimean-congo hemorrhagic fever virus(CCHFV) in Escherichia coli and obtain monoclonal antibodies(MABs) against NP protein.Methods:The recombinant NP protein was expressed in E.coli and purified.BALB/C mice were immunized with recombinant NP Protein.Hybridoma cell lines secreting MABs against NP protein were screened by regular cell fusion and indirect ELISA approach.The specificities of these MABs were determined by Western blot and immunofluorescecence assay.Results:Two hybridoma cell lines(10B12 and 3G5) stable in secreting specific MABs were successfully obtained.Western blot showed that two MABs from mouse ascites could bind specifically to NP protein.Indirect immunofluorescence assay indicated that they could specifically bind to NP protein expressed in the recombinant baculovirus and the specific fluorescene could be observed in infected insect cells.ELISA with 10B12 MAB as the first antibody and HRP-3G5 MAB as the second antibody showed that the linear range of the detected NP protein was from 1 to 20 ng.Conclusion:MABs of high specificity against NP protein have been successfully prepared,which lay the foundation for colloidal gold immunoassay of diagnosing CCHFV.
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关键词
NP protein,Monoclonal antibody,Crimean-congo hemorrhagic fever virus
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