布鲁氏菌抗体胶体金免疫层析法快速检测技术的建立

目的建立一种快速检测布鲁氏菌抗体的新方法。方法利用胶体金免疫层析技术,以大肠埃希菌表达、纯化的OMP31与BP26重组蛋白作为检测抗原,以金黄葡萄球菌A蛋白(SPA)作为胶体金标记物,制备胶体金免疫层析试纸条,用于检测布鲁氏菌抗体,并分析方法的敏感性和特异性。结果以粒径为40nm胶体金制备的试纸条检测布鲁氏菌抗体的敏感性最高,胶体金最佳标记pH为6.2,SPA蛋白最适标记量为6μg/ml。交叉试验证明试纸条不与其他非相关疾病感染血清反应,特异性高。试纸条检测结果与琥红平板试验方法的符合率为92%。结论制备的布鲁氏菌抗体胶体金免疫层析试纸条具有敏感、特异、简便、快速的特点,可用于鉴别布鲁氏菌自然感染和人工免疫,并可区分牛、羊种布鲁氏菌感染,可在基层推广使用。

Establishment of a gold immunochromatographic assay for rapid detection of Brucella antibodies

Objective To establish a new method of rapidly detecting Brucegga antibodies. Methods The two recombinant proteins BP26 and OMP31 were expressed in Eseherichia coli and purified to serve as detection antigens and a colloidal gold solu tion was conjugated with staphylococcal protein A （SPA） to provide an SPA-colloidal gold conjugate. These components were used to prepare colloidal gold immunochromatographic test strips for a gold immunochromatographic assay （GICA） to detect Bru- cella antibodies. The sensitivity and specificity of the assay were then analyzed. Results Strips prepared with 40-nm particles of colloidal gold were found to have the highest sensitivity. The optimal pH for labeling with colloidal gold was 6.2. The optimal amount of SPA for colloidal gold labelling was 6 μg/ml. Cros〉reactivity testing indicated that the strips did not react with sera positive for other unrelated diseases and that the strips were highly specific. The rate of concordance between strips and the Rose Bengal plate test （RBPT） was 92%. Conclusion Colloidal gold immunochromatographic test strips were prepared to detect Brucella antibodies. These strips are sensitive, specific, convenient, and quick. These strips can he used to identify natural Bru- cella infection and artificial immunity and distinguish between Brucella abortus infeetion and Brucella melitensis infection. These strips can be widely used at the basic level.