基于金纳米球的肠炎沙门菌SERS免疫层析快速检测技术

Gold nanoparticles-based surface-enhanced Raman scattering immunochromato⁃graphy for the rapid and quantitative detection of Salmonella Enteritis

为实现肠炎沙门菌的快速定量分析,采用Tris辅助柠檬酸钠还原法制备新型胶体金并利用Au-S配位作用在纳米金球表面修饰一层4-巯基苯甲酸(4-MBA)拉曼信号分子,然后以此为标记材料建立肠炎沙门菌表面增强拉曼(surface-enhancement of Raman scattering,SERS)免疫层析检测技术,并对其灵敏度、特异性和重复性进行考察。结果表明:建立的SERS免疫层析技术对肠炎沙门菌的可视化最低检测限(LOD)为10^(6) CFU/mL,而通过拉曼信号检测时LOD为10^(5) CFU/mL,灵敏度比裸眼观察提高了10倍。在10^(5)~10^(8) CFU/mL的浓度范围可根据标准曲线y=30.23×exp(-x/-147)-215.83(R^(2)=0.995)对肠炎沙门菌进行定量分析。该SERS免疫层析技术与鼠伤寒沙门菌、鸡白痢沙门菌、甲型副伤寒沙门菌等多种血清型沙门菌和大肠杆菌、副溶血弧菌等常见病原无交叉反应,特异性良好。同一批次的不同试纸条和不同批次的试纸条对相同样品的检测结果基本一致,重复性良好。以粪便和鸡蛋壳为基质的加标试验回收率在94.7%~104.9%之间,回收率良好。结果表明,基于金纳米颗粒的肠炎沙门菌SERS免疫层析快速检测技术灵敏度较高,特异性强,重复性好,能弥补传统胶体金试纸条无法定量的不足,可用于肠炎沙门菌的快速定量检测。

To realize the rapid and quantitative detection of Salmonella Enteritis,a novel colloidal gold was prepared using the Tris-assisted sodium citrate reduction method.A layer of 4-mercaptobenzoic acid(4-MBA)Raman signal molecule was conjugated on the surface of the gold nanoparticles via Au-S coordination,which was then used as the labeling material for establishing a SERS immunochromatographic assay of S.Enteritis.Finally,the sensitivity,specificity and reproducibility of the assay were evaluated.The results showed that the established SERS immunochromatographic technique had a visual limit of detection(LOD)of 10^(6) CFU/mL for S.Enteritis,while the LOD was 10^(5) CFU/mL according to the Raman signal intensity,which was 10-fold higher than that based on naked eye observation.Quantitative analysis of S.Enteritis could be performed in the concentration range of 10^(5)-10^(8) CFU/mL based on the standard curve of y=30.23×exp(-x/-147)-215.83(R^(2)=0.995).This SERS immunochromatographic technique had no cross-reactivity with other Salmonella serotypes such as S.Typhimurium,S.Pullorum,S.Paratyphi A,and other common pathogens such as Escherichia coli and Vibrio parahaemolyticus,show-ing a strong specificity.The results of different test strips from the same batch and different batches for the same samples were generally con-sistent,suggesting good reproducibility.The recoveries of the spiked feces and egg samples ranged from 94.7%to 104.9%,showing good re-coveries.The above results suggested that this gold nanoparticle-based SERS immunochromatographic technique with high sensitivity,high specificity and good repeatability,could be used for the rapid and quantitative detection of S.Enteritis.