摘要
为制备非洲猪瘟病毒(ASFV)MGF360-9L蛋白的多克隆抗体并评价其对病毒复制的影响,使用DNAStar 2.1软件预测MGF360-9L的抗原表位,选择并合成了符合抗原要求的MGF360-9L C端多肽CKNLSIAHKHYINDGFND,制备MGF360-9L家兔源多克隆抗体。随后采用实时荧光定量PCR方法、血细胞吸附试验和Western-blot方法等评价MGF360-9L的多克隆抗体对病毒复制的影响。结果显示,不同稀释倍数的MGF360-9L多克隆抗体在不同感染时间下均可显著降低病毒的拷贝数、效价以及D1133L和B646L(p72)两种病毒蛋白的表达,并且抗体稀释度越低,降低病毒拷贝数、效价以及病毒蛋白表达的作用越强。此外,不同稀释倍数的MGF360-9L多克隆抗体均可抑制感染重组病毒ASFV-GFP的猪肺泡巨噬细胞中绿色荧光蛋白的产生。结果表明,与阴性家兔血清相比,不同稀释倍数的MGF360-9L抗体均可显著抑制ASFV的复制。本研究为ASFV靶向药物和疫苗的研发提供了参考依据。
To prepare polyclonal antibodies against African swine fever virus(ASFV)MGF360-9 L protein and evaluate its effect on virus replication,the antigenic epitope of MGF360-9 L was predicted by DNAStar 2.1 software,and the MGF360-9 L C-terminal polypeptide C-KNLSIAHKHYINDGFND was selected and synthesized to prepare MGF360-9 L rabbit polyclonal antibody.Then the effect of polyclonal antibody against MGF360-9 L on virus replication was evaluated by real-time quantitative PCR,50%hemadsorpting doses assay and Western-blot method.The results showed that MGF360-9 L polyclonal antibody of different dilution times could significantly reduce the virus copy,titer and the expression of D1133 L and B646 L(p72)protein in different infection time,and the lower the antibody dilution,the stronger the effect of reducing virus copy,titer and expression of virus protein.In addition,MGF360-9 L polyclonal antibody of different dilution times could inhibit the production of green fluorescence protein in porcine alveolar macrophages infected with recombinant virus ASFV-GFP.The results suggest that MGF360-9 L polyclonal antibody of different dilution times could significantly inhibit the replication of ASFV compared with negative rabbit serum.This study provides a reference basis for the research and development of targeted drugs and vaccines for ASFV.
作者
杨博
申超超
张婷
郝雨
杨金柯
崔卉梅
赵登率
袁兴国
陈学辉
鲍恩东
张克山
郑海学
刘湘涛
YANG Bo;SHEN Chao-chao;ZHANG Ting;HAO Yu;YANG Jin-ke;CUI Hui-mei;ZHAO Deng-shuai;YUAN Xing-guo;CHEN Xue-hui;BAO En-dong;ZHANG Ke-shan;ZHENG Hai-xue;LIU Xiang-tao(College of Veterinary Medicine,Nanjing Agricultural University,Nanjing 210095,China;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)
出处
《中国兽医科学》
CAS
CSCD
北大核心
2022年第3期324-332,共9页
Chinese Veterinary Science
基金
甘肃省科技重大专项(20ZD7NA006)
国家自然科学基金联合项目(31941002)
国家重点研发计划项目(2018YFC0840400)。
