基于智能水凝胶的适体-压电生物传感器检测大肠杆菌O157:H7

The development of new methods for rapid detection of foodborne pathogenic bacteria remains a challenging and important issue for ensuring food safety and security. A new piezoelectric aptasensor based on intelligent hydrogel will be developed for the detection of Escherichia coli O157:H7 in this proposal. The overall goal of the proposed project is to develop a unique material, ssDNA and aptamer crosslinked polyacrylamide hydrogel which is specifically responsive to Escherichia coli O157:H7 and to demonstrate the feasibility for fabrication of piezoelectric aptasensor coated with the intelligent hydrogel to be developed. The core of the proposed novel hydrogel is to introduce deoxyribonucleic acid (ssDNA) and aptamer against Escherichia coli O157:H7 to the polymer backbones to form a 'smart' Escherichia coli O157:H7-responsive hydrogel. The specific research will be: 1). Synthesis and optimization of polyacrylamide with ssDNA and aptamer as crosslinker; 2). Fabrication of the film piezoelectric aptasensor with synthesized aptamer hydrogels; 3). Detection of killed Escherichia coli O157:H7 in buffer solution using developed polymer hydrogel coated piezoelectric aptasensor. The research will prove the feasibility of ssDNA and aptamer crosslinked polyacrylamide hydrogel coated piezoelectric aptasensor for rapid detection of killed Escherichia coli O157:H7. The proposed piezoelectric aptasensor will be a new alternative approach and significant promotion in the field of piezoelectric biosensor for the detection of pathogenic bacteria.

用于致病菌的快速检测技术是遏制食源性疾病暴发，加强食品安全的保障，具有重大的社会和经济意义。本项目以大肠杆菌O157:H7为研究对象，在设计、筛选对目标具有特异性响应适体的基础上，合成一种独特的以ssDNA和适体作为交联剂的聚丙烯酰胺水凝胶，能对目标作出特异性响应；进而将该水凝胶修饰于传感器表面，开发基于智能水凝胶的适体-压电生物传感器，并对大肠杆菌O157:H7标准液进行检测，评估此适体-压电生物传感器检测目标的性能。本项目选题研究是对压电生物传感器检测致病菌领域的全新突破，是对抗体-压电生物传感器检测致病菌的挑战和创新性研究，对生物传感器检测致病菌方法的发展具有重大的理论和实际意义。

直接、快速、灵敏的检测方法一直是食品安全检测领域面临的挑战。本项目瞄准这一需求，研究合成了一种以ssDNA和适体作为交联剂的聚丙烯酰胺智能水凝胶，并在其基础上构建了基于智能水凝胶的压电适体传感器，建立了一种能够对目标实现直接、快速检测分析的方法。项目以凝血酶标准液为试验对象，验证了基于水凝胶构建的压电适体传感器对目标进行快速检测分析的可行性；建立了传感器频率响应与目标浓度之间的相关性；并用AFM对传感器进行了表征。项目研究表明：丙烯酰胺、TBA29、ssDNA三种单体以10:1:1的比例合成水凝胶，构建的适体传感器灵敏度最高，在30分钟内能够直接检测到1nM凝血酶。与未采用该水凝胶构建的压电适体传感器相比，其灵敏度提高了一个数量级。本项目的研究为生物传感器快速检测分析领域提供了一种新方法，其研究成果在食品安全检测领域具有重要的理论和实际意义。