斑点免疫金渗滤法检测赤羽病抗体的研究

以亲和层析原理为基础,将提纯的AKAV抗原包被在硝酸纤维膜上,利用胶体金标记SPA显色,建立了斑点免疫金渗滤法诊断赤羽病抗体的方法。其中最佳点样抗原质量浓度是0.099mg/mL,封闭液选择含1%BSA,0.5%Tween-20的0.01mol/LpH7.2的PBS缓冲液,SPA胶体金最佳稀释度为1∶4。特异性试验表明,该方法特异性好,与牛病毒性腹泻病毒、牛传染性鼻气管炎、牛白血病、口蹄疫、蓝舌病等阳性血清不发生交叉反应。将DIGFA与ELISA进行对比,差异不显著。研究结果表明,该方法操作简单,结果易于判断,适用于赤羽病的临床诊断和流行病学调查。

Dot Immunogold Filtration Assay for Detection of Antibodyagainst Akabane Virus

Based on the affinity chromatography,a new dot immunogold filtration assay（DIGFA）was established to detect serum antibody against Akabane virus.The purified antigens of AKAV were coated on nitrocellulose（NC）membrane,and staphylococcal protein A（SPA）labeled with colloidal gold was used for reaction reagent.The optimum Ag concentration was 0.099 mg/mL.The optimal blocking buffer was 0.01mol/L,pH7.2 PBS containing 1%BSA and 0.5% tween-20.The optimal dilution of SPA labeled with colloidal gold was 1∶4.It was found that the specificity of this assay was high for AKAV without any cross-reactions with other viruses positive sera,such as BVDV、IBRV、BLV、FMDV 、PRV,and so on.Compared DIGFA and ELISA,it was found that the two results were not remarkable different.It suggested that the method is easily and quickly determined and very useful for clinical diagnosis and epidemiological survey of AKA.