CAJ | 학술논문

目的：建立一种基于表面等离子共振技术(Surface Plasmon Resonance technology, SPR)原理的生物传感器方法,实现11种常见致病菌的高通量检测。方法用表面自组装技术(self-assembled monolayer, SAM)在金膜表面引入羧基,然后通过氨-羧基反应将5’端标记有氨基的探针固定在芯片表面,经多重PCR扩增出的产物变性、杂交、生物素-亲和素系统信号放大等步骤完成目的片段的检测。对所建立的多重PCR-SPR检测方法的灵敏性、特异性等技术指标进行评价。结果本研究建立的多重PCR-SPR检测方法具有良好的灵敏性,该芯片检测11种常见致病菌的灵敏性均达到103~104 cfu/mL,能满足现有检测要求。通过对实验室保存的14株标准菌株和沙门氏菌等90份样品分离株进行检测,结果显示该检测方法具有较好的特异性,无交叉反应。结论本研究建立的多重 PCR-SPR 检测系统具有良好的灵敏性和特异性,完全适用于11种致病菌的高通量检测。该检测方法检测时间短、检测成本低、减少了人为因素的影响,具有良好的应用前景。
목적：건립일충기우표면등리자공진기술(Surface Plasmon Resonance technology, SPR)원리적생물전감기방법,실현11충상견치병균적고통량검측。방법용표면자조장기술(self-assembled monolayer, SAM)재금막표면인입최기,연후통과안-최기반응장5’단표기유안기적탐침고정재심편표면,경다중PCR확증출적산물변성、잡교、생물소-친화소계통신호방대등보취완성목적편단적검측。대소건립적다중PCR-SPR검측방법적령민성、특이성등기술지표진행평개。결과본연구건립적다중PCR-SPR검측방법구유량호적령민성,해심편검측11충상견치병균적령민성균체도103~104 cfu/mL,능만족현유검측요구。통과대실험실보존적14주표준균주화사문씨균등90빈양품분리주진행검측,결과현시해검측방법구유교호적특이성,무교차반응。결론본연구건립적다중 PCR-SPR 검측계통구유량호적령민성화특이성,완전괄용우11충치병균적고통량검측。해검측방법검측시간단、검측성본저、감소료인위인소적영향,구유량호적응용전경。
Objective A biosensor based on surface plasmon resonance (SPR) principle was developed for food-borne pathogen bacteria detection, the method achieved a high-throughput detection for 11 kinds of bacteria.Methods The chip surface was modified by self-assembled monolayer (SAM), and carboxyl group was introduced. The reaction of the carboxyl and amino leading to the probes were arrayed on the chips, and the biotin-avidin system was introduced to enhance the signal after hybridization. We evaluated the sensitivity, the specificity and other technical indicators by multiple PCR-SPR method.Results The multiplex PCR-SPR method showed a good sensitivity. The sensitivity of 11 kinds pathogenic bacteria reached 103~104 cfu/mL, which can meet the requirements of existing detection. Detection of 14 Standard strains and Salmonella strains were isolated from 90 samples. The results showed that the method had a good specificity without cross reaction.Conclusion In this study, a detection method for 11 kinds of common foodborne pathogens was es-tablished by multiplex PCR-SPR system. It has a short detection time, a low detection cost, a reduction of the influence of manual factors, and will have a good application prospect.