Establishment of an Indirect ELISA for Detecting Egg Yolk Antibody Against Chicken Flavivirus
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摘要: 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。Abstract: 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 ·μL-1,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 substrate,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.
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Key words:
- chicken-origin Flavivirus /
- indirect ELISA /
- egg yolk antibody
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