Yuan Jinglei, Wu Shijia, Duan Nuo, Ma Xiaoyuan, Xia Yu, Chen Jie, Ding Zhansheng, Wang Zhouping
State Key Laboratory of Food Science and Technology, Synergetic Innovation Center of Food Safety and Nutrition, School of Food Science and Technology, Jiangnan University, Wuxi 214122, China.
State Key Laboratory of Food Science and Technology, Synergetic Innovation Center of Food Safety and Nutrition, School of Food Science and Technology, Jiangnan University, Wuxi 214122, China.
Talanta. 2014 Sep;127:163-8. doi: 10.1016/j.talanta.2014.04.013. Epub 2014 Apr 13.
In this study, a gold nanoparticle-based colorimetric aptasensor for Staphylococcus aureus (S. aureus) using tyramine signal amplification (TSA) technology has been developed. First, the biotinylated aptamer specific for S. aureus was immobilized on the surface of the wells of the microtiter plate via biotin-avidin binding. Then, the target bacteria (S. aureus), biotinylated-aptamer-streptavidin-HRP conjugates, biotinylated tyramine, hydrogen peroxide and avidin-catalase were successively introduced into the wells of the microtiter plate. After that, the existing catalase consumed the hydrogen peroxide. Finally, the freshly prepared gold (III) chloride trihydrate was added, the color of the reaction production would be changed and the absorbance at 550 nm could be measured with a plate reader. Under optimized conditions, there was a linear relationship between the absorbance at 550 nm and the concentration of S. aureus over the range from 10 to 10(6) cfu mL(-1) (with an R² of 0.9947). The limit of the developed method was determined to be 9 cfu mL(-1).
在本研究中,已开发出一种基于金纳米颗粒的比色适体传感器,用于金黄色葡萄球菌检测,该传感器采用了酪胺信号放大(TSA)技术。首先,通过生物素-抗生物素蛋白结合将对金黄色葡萄球菌具有特异性的生物素化适体固定在微量滴定板孔的表面。然后,将目标细菌(金黄色葡萄球菌)、生物素化适体-链霉亲和素-辣根过氧化物酶缀合物、生物素化酪胺、过氧化氢和抗生物素蛋白-过氧化氢酶依次引入微量滴定板的孔中。之后,现有的过氧化氢酶消耗过氧化氢。最后,加入新制备的三水合氯化金(III),反应产物的颜色会发生变化,并用酶标仪测量550nm处的吸光度。在优化条件下,550nm处的吸光度与金黄色葡萄球菌浓度在10至10(6) cfu mL(-1)范围内呈线性关系(R²为0.9947)。所开发方法的检测限确定为9 cfu mL(-1)。