基于喹诺酮类双识别仿生抗体的表面等离子体共振传感检测方法及识别机制研究

In animal-derived foods, quinolone residues can cause the human body to produce a variety of toxic responses. Although the residues can be detected rapidly by the antibody-based biosensor, the antibody has the limitations of long-period preparation, instability, high cost and inconvenient storage. It is urgent to develop a new biomimetic antibody with the characteristics of simple preparation, stable properties and high specificity. Based on the accurate and precise recognition model of biomacromolecules, this project plans to develop a biomimetic antibody with double recognition for aiming at the problems of the existing antibody. Combined with molecular imprinting technology, the recognizing specificity will be improved by means of quinolone-specific aptamers. A quinolone surface plasmon resonance sensing method will be developed by applying chemical modifications and sensitization enhancement by nano-scale gold particles. By characterizing the physical and chemical properties and recognition performance, the structure-activity relationship and influencing factors will be analyzed, from which the law of bionic design and the mechanism of interaction can be explored. The constructing model of the new sensor will be investigated after clarifying the sensing recognition mechanism. In this project, the specific aptamer screening method is combined with molecular imprinting technology. The biologically recognizing principle will be deduced and verified using chemical methods. The project represents a form of interdisciplinarity. It will be contributed to break down and solve the difficult problems of molecular imprinted polymers with low specificity and aptamers with poor stability. The implementation of this project will provide new thoughts for the design of high-performance molecularly imprinted polymers and technical supports for monitoring residues of harmful substances in foods.

动物性食品中喹诺酮类残留能使人体产生多种毒性反应，基于抗体的生物传感器虽然实现了该类药物残留的快速检测，但抗体制备周期长、不稳定、成本高且不便储存,亟需研发便于制备、性质稳定和特异性强的新型仿生抗体。本项目针对已有抗体存在的问题，基于生物大分子的精准识别模式，借助喹诺酮类特异性适配体，结合分子印迹技术，研制双识别仿生抗体，提高识别的专一性，运用化学修饰与纳米金增敏技术，建立喹诺酮类表面等离子体共振传感检测方法。通过表征仿生抗体的理化性质和识别性能，分析构效关系和影响因素，揭示仿生设计规律和相互作用机理，阐明传感识别机制，探索新型传感器的构建模式。本研究将特异性适配体筛选方法与分子印迹技术相结合，应用化学方法模拟和推演生物学识别原理，体现了学科交叉，有助于破解分子印迹聚合物特异性差、适配体稳定性不足的难题。项目的开展将为高性能仿生抗体的设计提供新思路，为食品中有害物质残留的监控提供技术支撑。

喹诺酮类药物具有抗菌效果好、组织穿透力强和价格低廉等优点，广泛用于动物和人类感染性疾病的防治，随着该类药物滥用和误用现象的不断增多，在动物性食品中的残留日益严重，由此引发的食品安全风险显著增高，现有检测方法在特异性、稳定性和经济性等方面仍亟待改进。在前期研究基础上，本项目借鉴生物大分子的精准识别模式，应用分子印迹技术，结合特异性适配体和生物源性交联功能单体，研制双识别仿生印迹抗体，建立喹诺酮类分子印迹表面等离子体共振传感检测方法。该方法对实际样品中喹诺酮类药物检测的线性范围为50 ng/L-2000 ng/L，线性相关系数为0.9995-0.9999，检测限为7.41 ng/kg-21.34 ng/kg，定量限为19.22 ng/kg-64.74 ng/kg，样品的回收率为75.21%-105.17%，批内变异系数为15.33%-25.24%，批间变异系数为23.75%-34.11%。本研究通过理化性质和识别性能表征，明确了高性能人工抗体构建的技术优势和发展潜力，总结了仿生印迹规律，分析了相互作用机理，探索了传感识别模式。项目的开展，有助于充实分子印迹技术的理论内涵，推进不同学科研究方法和技术手段的交叉融合，建立并发展新型分析方法和监测手段，确保食品安全和人类健康。在本项目的资助下，以第一作者发表中国科学院一区SCI/TOP期刊论文1篇，以通讯作者发表SCI、EI和中文核心刊物论文各1篇，参与发表SCI论文1篇；做特邀学术报告1次；获省级和地厅级优秀科技论文一等奖各1项；荣获相关荣誉称号6项；培养硕士研究生7名，本科生15名。