汉滩病毒核壳蛋白（NP）在杆状病毒系统的表达及其免疫原性研究

BACULOVIRUS EXPRESSION OF THE SMALL GENOME SEGMENT(S)OF HANTAAN VIRUS AND THE IMMUNOGENICSTUDIES ON THE EXPRESSED NUCLEOCAPSID PROTEIN(NP)

应用杆状病毒表达载体成功地表达了汉滩病毒７６－１１８株（ＨＴＮＶ）核壳蛋白。将ＨＴＮＶＳ基因插入杆状病毒转染质粒ｐＡｃＹＭＩＢ的多角体基因启动子下游附近，与经Ｂｓｕ３６Ｉ酶切线性化的杆状病毒（ＡｃｖＥＰＡ）ＤＮＡ共同转染Ｓｆ９细胞，经空斑筛选获得了高效表达ＮＰ的重组杆状病毒（ＡｃＶＨａｎＳ）。经ＳＤＳ－ＰＡＧＥ和Ｗｅｓｔｅｒｎｂｌｏｔ证实，表达产物与ＨＴＮＶ毒粒ＮＰ分子量均为５０ｋＤ左右，紫外扫描显示表达量约占细胞总蛋白量的１２．１％，ＥＬＩＳＡ滴度达１：３２００～１：６４００。用一批抗ＮＰ单克隆抗体和病人血清证实，重组ＮＰ与毒粒ＮＰ的免疫反应性完全一致。

he recombinant baculovirus, AcVHanS,which contains the S segement of Hantaan Virus was generated by cotransfection of Sf9 cells with the recombinant plasmid (pAcHanS)andBsu36Ⅰlinearized AcVEPA DNA.A high level expression of the HTNV NP was obtained with the baculovirus recombinant.The expressed NP was electrophoretically identified and Wasindistinguishable from the authentic viral NP in molecular weight. Its identity was further con-firmed by Western blot.The expressed NP constituted about 12.1%of total proteins of AcVHanS infected Sf9 cells.The antigen titres by ELISA was 1:3,200-1:6,400.The reactivitieswith HFRS patients’and a panel of MAbs against NP indicated the identity of the ex- pressed NP with the authentic viral NP.