【摘要】 采用活病毒免疫,常规杂交瘤融合的方法,获得了4株针对H5N1亚型禽流感病毒(AIV)A/duck/Shandong/093/2004(A/D/SD/04)株血凝素(HA)的单克隆抗体A8E8、D2F8、A8C8和C3G4。其中A8E8可以与11株不同来源的H5N1AIV发生交叉免疫抑制性反应(HI),具有广谱性。采用间接免疫荧光试验(IFA),用A8E8单克隆抗体测定A/D/SD/04株对COS1和MDCK2种细胞的半数感染量(TCID50),结果显示,A/D/SD/04株在COS1上的TCID50为10-5.33/0.1mL,在MDCK上的为10-7.33/0.1mL。将编码A/D/SD/04株的HA基因质粒与PR8质粒共转染COS1细胞后,用A8E8进行IFA,能观察到特异性绿色荧光,这与鸡胚接种结果相符。表明,用此单克隆抗体能准确地检测到H5重组病毒。更多还原

【Abstract】 The BALB/c mice were immunized with active virulent, H5N1 subtype avian influenza virus strain A/duck/Shandong/093/2004(A/D/SD/04), and fusion of lymphocyte and myeloma SP2/0 cell was carried out by conventional method. Four monoclonal antibodies against hemagglutinin protein of H5N1 subtype avian influenza virus were obtained, which were designated as A8E8,D2F8,A8C8 and C3G4 respectively. A8E8 could reacted with 11 strains of H5N1 subtype avian influenza virus isolated from ~chicken , duck, goose and quail in different places from 2001 to 2004. CID_~50 of A/D/SD/04 was detected in MDCK and COS-1 cells by using indirect immunofluorescence assay (IFA). The result showed that the virus could grow well in COS-1 with the CID_~50 10~-5.33 /0.1mL, and the cell was suitable to rescue of A/D/SD/04. PHW-HA of A/D/SD/04 and 7 plasmids of PR/8 strain were cotransfected into COS-1 cell and recombinated viruses were produced after inoculating the supernatant into SPF embryonated chicken eggs, which was confirmed by IFA with A8E8. These results suggested that the four monoclonal antibodies will be extremely useful in detection of A/D/SD/04.更多还原