鸡源黄病毒卵黄抗体间接ELISA检测方法的建立

Establishment of an Indirect ELISA for Detecting Egg Yolk Antibody Against Chicken Flavivirus

为在检测产蛋鸡黄病毒抗体水平过程中减少应激,以鸡源黄病毒FQ-C1株为包被抗原,建立检测鸡源黄病毒卵黄抗体的间接ELISA方法。确定的优化条件是包被抗原浓度为5.7ng.μL-1,卵黄抗体以1∶160倍稀释,羊抗鸡IgG酶标抗体以1∶3 200稀释。经特异性、敏感性检验表明该方法特异性强、敏感性高,适合于产蛋鸡免疫后的抗体检测。

A novel Flavivirus infection, which mainly caused egg-dropping in ducks and chickens, has spread throughout China since 2010. To carry out the serological surveillance in chickens with minimum stress, an indirect enzyme-linked immuosorbent assay （ELISA） for detecting egg yolk antibody against Flavivirus in layer chickens has been developed by coating plate with chicken-origin Flavivirus FQ-C1 isolate. The optimal working concentration of antigen was 5.7 ng ~ /~L ~, the working concentration of egg yolk antibody was 1 ： 160 dilution and that of HRP- labeled goat anti-chicken IgG was 1 ： 3 200. After a color reaction of 10 minutes with Tetramethylbenzidine （TMB） liquid suhstrate, a threshold of positive/negative （P/N） 〉 2. 1 was determined as positive standard for the ELISA. The results indicated that the ELISA established by this research provided a high sensitivity and specificity test, which was suitable for the detection of antibodies against Flavivirus in immune laying hens.