黄热病毒单克隆抗体的制备及免疫反应特性鉴定

目的 制备黄热病毒单克隆抗体并对其生物特性进行鉴定.方法 通过动物免疫、细胞融合、阳性克隆筛选等方法制备黄热病毒单克隆抗体,免疫Balb/c小鼠收集腹水,得到扩大制备的单抗,辛酸-饱和硫酸铵法纯化腹水.IFA法检测单抗与黄热病毒(YFV)、乙脑病毒(JEV)、登革病毒1-4型(DENV1-4)、西尼罗病毒(WNV)和基孔肯雅病毒(CHIKV)的特异性反应,并对纯化前后的腹水进行效价分析；ELISA方法进行单抗的亚型分类.结果 获得8株黄热病毒的单克隆抗体,均与YFV有特异性反应,1株与JEV有交叉反应,余7株与其它4种虫媒病毒无交叉反应.抗体分型大部分为IgG；纯化后单抗蛋白含量大多在30 mg/ml以上.纯化后的4株单抗效价在1∶25600,3株在1∶12800,仅1株效价较低.结论 本研究目前获得了几株特异性好、效价高的黄热病毒单克隆抗体,为建立黄热病毒快速检测系统奠定了基础.

Preparation and Characterization of the Monoclonal AntibodiesAgainst Yellow Fever Virus

Objective To prepare mouse monoclonal antibodies （McAbs） against yellow fever virus （YFV） and evaluate their biological characteristics. Methods McAbs against YFV were prepared by immunizing mouse, cells fusing, cloning and screening. McAbs were prepared from the ascites collected from hybirdoma clone injected BALB/c mice. The McAbs were purified by Octanoic acid-saturated ammonium sulfate method. The specificity of the MeAhs was check against YFV, Japanese encephalitis virus （JEV）, dengue virus 1-4 （DENVI-4）, west nile virus （WNV） and Chikungunya virus （CHIKV） by immunofluorescence assay （IFA）. McAbs isoforms were determined by ELISA. Results Eight McAbs showed specific binding to yellow fever virus but only one MeAb had a cross-reaction with JEV. Other seven MeAbs had no cross reactions with the other four strains of arboviruses.Typing results showed almost McAbs were IgG. Purified McAbs proteins had a concentration over 30 mg/ml. Four purified McAbs titers level were 1：25600 and three McAbs were 1：12800 except one with lower titer. Conclusion Several YFV McAbs were obtained with good specifieities and high titers which established foundation for quick detection system of Yellow Fever virus.