摘要
基于适配体识别和荧光探针技术,发展了一种检测肠致病大肠杆菌（EPEC）的新型分析方法。该方法以生物素化适配体1为捕获探针,通过生物素与亲和素的特异性结合得以固定在微孔板中;以FAM标记的适配体2作为显示探针,当有目标物存在的情况下,生物素化适配体1将捕获目标物质并与其相结合,同时FAM标记的适配体2也会与目标物质相结合,从而使得目标物标记上荧光信号,通过测定荧光信号可实现对目标物质的定量检测。考察了影响方法分析性能的各种条件,结果发现当微孔板包被亲和素质量浓度为100μg/m L,生物素化适配体1浓度为100 nmol/L,FAM标记的适配体2浓度为150 nmol/L时,可获得方法的最佳分析表现。在最佳试验条件下,方法对肠致病大肠杆菌检测的线性范围为50~106cfu/m L,检出限为50 cfu/m L。方法选择性良好,操作简单,已成功应用于实际样品中肠致病大肠杆菌的检测。
A novel analytical method for the detection of Enteropathogenic Escheriehia coli （EPEC） was established based on the aptamer recognition and fluorescent probe technology. Biotinylated aptamer 1 was used as capture probe, which was immobilized onto the microwell plate through the specific binding of biotin and avidin. Fluoreseein amidite （FAM） labeled aptamer 2 was used as report probe. In the presence of target, the biotinylated aptamer 1 would capture and bind with target. In addition, FAM labeled aptamer 2 would also bind with target and displayed fluorescence signal. All the condition factors affecting the performmance of the present method were investigated. The results show that when the avidin concentration coated on the microplate is 100 μg/mL, biotinylated aptamer 1 concentration is 100 nmol/L, and the concentration of FAM labeled aptamer 2 is 150 nmol/L, the optimal analytical performance can be achieved for the present work. Under the optimal conditions, the linear range for the EPEC concentration detection is 50-106 cfu/mL with a detection limit of 50 cfu/mL. Meanwhile, the present method is highly selective for OTA and easy to be operated. It has been successfully applied to measure EPEC content in real samples.
出处
《中国食品学报》
EI
CAS
CSCD
北大核心
2015年第12期160-165,共6页
Journal of Chinese Institute Of Food Science and Technology
基金
国家自然科学基金项目(31401575)
江苏省自然科学基金项目(BK20140155)
关键词
适配体
荧光分析
肠致病大肠杆菌
aptamer
fluorescence analysis
Enteropathogenic Escherichia coli
作者简介
段诺，女，1986年出生，博士，副教授
通讯作者：王周平
