布鲁氏菌荧光微球免疫层析试纸条的研制
Development of Fluorescence Microsphere Immunochromatographic Strips for Brucellosis
杨若松 1马增彬 1龚雅云 1蔡云虹 1南文龙 2李知新3
作者信息
- 1. 百沃特(天津)生物技术有限公司,天津 301700
- 2. 中国动物卫生与流行病学中心,山东青岛 266032;农业农村部反刍动物重大疫病防控重点实验室(东部),山东青岛 266032
- 3. 宁夏回族自治区动物疾病预防控制中心,宁夏银川 750000
- 折叠
摘要
本研究以量子点荧光微球为标记物,制备免疫层析试纸条,用于快速检测牛血清布鲁氏菌抗体.采用布鲁氏菌脂多糖(LPS)和兔抗重组链球菌蛋白G多克隆抗体分别作为检测线和质控线喷涂硝酸纤维素膜,以荧光微球标记的重组链球菌蛋白G喷涂玻璃纤维素膜作为量子点结合垫,经组装、切割制备免疫层析试纸条.使用布鲁氏菌病阳性血清国家标准品测定所建立方法的灵敏度和稳定性,使用牛鼻气管炎阳性血清等常见牛病血清及牛布鲁氏菌病阴性血清国家标准品评价该方法的特异性;使用建立的方法与间接ELISA(iELISA)同步检测 100 份临床血清样品,比较两者的符合率.结果显示:量子点荧光微球免疫层析试纸条的最低检出限为 0.3 IU/mL,常温保存 18 个月其检测灵敏度不变;与牛鼻气管炎、牛流行性腹泻、牛结节性皮肤病等临床疫病的血清抗体无交叉反应;与iELISA方法的阳性符合率为 94.2%,阴性符合率为 100%,总符合率为 97.0%.结果表明,本研究建立的布鲁氏菌荧光微球免疫层析试纸条敏感、特异、稳定,操作时不需要借助仪器,适用于临床快速检测和初筛,可为我国布鲁氏菌病防控提供技术支持.
Abstract
In this study,immunochromatographic strips were prepared taking quantum dot fluorescent microspheres as markers for rapid detection of bovine serum antibodies against Brucella.Brucella lipopolysaccharide(LPS)and rabbit polyclonal antibodies against recombinant streptococcal protein G were sprayed onto nitrocellulose membranes as detection and quality control lines,respectively.Recombinant streptococcal protein G labeled with fluorescent microspheres were sprayed onto glass cellulose membranes as quantum dot binding pads.After assembly and cutting,the immunochromatographic strips were prepared.The sensitivity and stability of the prepared strips were determined using national standard brucellosis positive serum,and the specificity was determined using common bovine disease serum including bovine rhinotracheitis positive serum and national standard brucellosis negative serum.100 clinical serum samples were synchronously tested using the prepared strips and indirect ELISA(iELISA)to compare the compliance rate of the two methods.The results showed that the lowest detection limit of the prepared strips was 0.3 IU/mL,and the sensitivity remained unchanged after storage for 18 months under room temperature;no cross reaction with negative serum of brucellosis or positive serum of bovine rhinotracheitis,bovine epidemic diarrhea and lumpy skin disease was found;compared with iELISA,the positive,negative and overall compliance rates were 94.2%,100%and 97.0%,respectively.In conclusion,the prepared strips were sensitive,specific and stable for rapid clinical detection without aid of instruments,supporting the prevention and control of brucellosis in China.
关键词
布鲁氏菌/荧光微球/免疫层析技术/脂多糖Key words
Brucella/LPS/fluorescent microsphere/immunochromatographic technology/lipopolysaccharide引用本文复制引用
基金项目
河北省重点研发计划(21327601D)
国家现代奶牛产业技术体系建设项目(CARS-36)
出版年
2024
国家科技期刊平台
NSTL
万方数据