迟缓爱德华氏菌胶体金快速检测试纸的研制

A Gold Immunochromatography Assay for the Detection of Edwardsiella tarda

本研究用纳米胶体金颗粒标记抗迟缓爱德华氏菌(AL60306NA1)单克隆抗体3A7并制备金标垫,将抗迟缓爱德华氏菌(AL60306NA1)单克隆抗体4F11与羊抗鼠抗体作为捕获抗体包被硝酸纤维素膜,建立迟缓爱德华氏菌的胶体金免疫层析快速检测方法。经过对试纸条的特异性和灵敏度测定,结果表明:试纸条与嗜水气单胞菌、温和气单胞菌、豚鼠气单胞菌、海安气单胞菌、产碱假单胞菌、无乳链球菌、大肠埃希杆菌、哈维氏弧菌、鳗弧菌、创伤弧菌、副溶血弧菌、类志贺邻单胞菌、鲁氏不动杆菌等15种水产常见病原菌没有交叉反应,与迟缓爱德华氏菌特异性反应,检测灵敏度为1×105cfu.mL-1,检测所需时间低于20min。所制备的迟缓爱德华氏菌胶体金免疫层析检测试纸条具有快速、简便、特异性高和适应基层生产推广应用等优点。

A rapid, accurate and easy gold immunochromatographic lateral flow assay system （GICA） was developed for detection of Edwardsiella tarda. The monoclonal antibody 3A7 against Edwardsiella tarda which was conjugated with colloidal gold acts as signal generator on conjugate pad. The monoclonal antibody 4Fll was used as a capture antibody at the test line （T）, goat anti-mouse IgG antibody was used as the capture antibody at the control line （C） on the nitrocellose membrane. We assembled the GICA test trip with absorbing pad, conjugate pad, nitrocellose membrane which spray with capture antibody. The preliminary feasibility study of this method was described as following： The sensitivity of the GICA test strip towards Edwardsiella tarda was high with a detection limit of 1 ×10^5cfu·mL^-1. The specificity of the assay showed no cross-reaction with other thirteen bacteria of aquaculture pathogens such as A. carla, A. sobria, Streptococcus agalactiae, et al. ; Accurate reading time for confirmation of the assay can be completed in 20 min with a liquid sample of 100 μL. This study indicated that the GICA test strip pro- vided high sensitivity and specificity for the detection of Edwardsiella tarda.