摘要
以禽流感病毒(AIV)湖北分离株(H9N2亚型)提纯物为免疫原,制备出鸡抗AIV和兔抗AIV抗体,经用琼脂扩散法测得效价分别为1∶32和1∶16。以纯化的鸡抗AIVIgG为包被抗体,兔抗AIVIgG为第二抗体,建立了检测AIV抗原的夹心ELISA法。结果表明,鸡抗AIVIgG的最佳包被浓度为1μg/mL,兔抗AIVIgG的最适工作浓度为5μg/mL;对已知的阳性样品,用夹心ELISA法测得的病毒滴度比血球凝集滴度高16倍以上,且能检出其它亚型的禽流感病毒;与新城疫病毒、传染性支气管炎病毒、减蛋综合征病毒、传染性喉气管炎病毒、传染性法氏囊病病毒、鸡痘病毒、马立克氏病病毒等无交叉反应,说明本方法有很高的特异性及敏感性。对14个鸡场送检的、患有呼吸道疾病或有腹膜炎、眼炎、产蛋下降、怀疑为禽流感感染的病鸡进行了检测,结果有7个鸡场为阳性。本方法的建立为禽流感病鸡群的临床检验提供了一种方便、敏感、快速的检测方法。
Chickens and rabbits were immunized with purified avian influenza virus (AIV) H_9N_2 subtype isolated from Hubei province. Antibody titers of chicken anti-AIV serum and rabbit anti-AIV serum in (agar) diffusion test were 1∶32 and 1∶16, respectively. A sandwich ELISA for detection of AIV antigen was developed by using purified chicken anti-AIV IgG coated on the ELISA plate and rabbit anti-AIV IgG as the second antibody. The results showed that the optimum working concentrations of chicken anti-AIV IgG and rabbits anti-AIV IgG were 1 μg/mL and 5 μg/mL, respectively. The dilution titers of positive AIV sample by ELISA was 16 times higher than that by hemagglutination test, and other AIV subtypes can be detected by sandwich ELISA. No cross reaction was observed with Newcastle disease virus (NDV), infectious bronchitis virus (IBV), eggs drop syndrome virus (EDS_(-76)V), infectious laryngotracheitis virus (ILTV), infectious bursal disease virus (IBDV), avian pox virus (APV) and Marek's disease virus(MDV). 14 suspicious chicken groups showing respiratory tract syndrome or peritonitis, opthalmitis ,egg drop were detected by the sandwich ELISA, and of them, 7 groups were positive. All results showed that the sandwich ELISA was specific and sensitive. It is suggested that the sandwich ELISA is a convenient, sensitive and quick method to detect AIV antigen.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2004年第5期536-541,共6页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
湖北省科技厅重点项目(20002P0805)
