表面增强拉曼光谱技术在多元病原菌同时检测中的应用策略

Application Strategies of Surface-Enhanced Raman Spectroscopy in Simultaneous Detection of Multiple Pathogens

水、空气、食品、灰尘和排泄物中广泛存在食源性病原菌,由此引发的感染性疾病严重危害人类健康。因此,开发病原菌的快速检测方法尤为必要。由于实际样品中的病原菌往往共生存在,所以多元病原菌的同步灵敏检测是微生物检测领域的重点与难点。分子生物学和免疫组化分析技术都在此领域进行过一些尝试,但由于引物设计与抗体的局限性,这两种技术在实际应用中的效果并不十分理想。表面增强拉曼光谱(SERS)技术由于具有快速、无损、高分辨率、不受水分干扰、可原位检测等显著优势,在多元病原菌同步检测领域获得了重要应用。从应用原理、特点和效果等方面出发,系统阐述了SERS技术在多元病原菌同时检测中的应用策略。首先对SERS基底材料与病原菌的结合方式进行简要概述,再以检测策略为主线,从直接法和间接法两种策略出发进行介绍。直接法通过基底材料的信号放大作用直接获得病原菌本身的光谱信息,步骤简便,操作快捷,在多元病原菌判别分析、定量分析与即时检测(POCT)中被广泛应用。但由于光谱信息量大,往往需要与多元统计分析方法、成像技术和微流控器件等联用。间接法一般需要借助拉曼信号分子和适配体、抗体等识别元件,将对病原菌的检测转换为对信号分子的分析,极大提高了检测方法的灵敏度与特异性,可在基因、蛋白、细胞等水平实现对多元病原菌的同步分析。且与其他识别元件及功能分子的联用能构建得到集细菌的分离、识别与灭活于一体的综合检测体系,在临床血液等实际样本的分析中具有重要前景。最后,总结并指出SERS技术的现有问题及下一步努力方向,为SERS技术在多元病原菌的快速、灵敏检测策略设计及具体应用方面提供参考。

Foodborne pathogens are widely present in water,air,food,dust,and excrement,the infectious diseases caused by these seriously endanger human health.Therefore,it is essential to develop rapid detection methods for pathogens.Since pathogens in actual samples often co-exist,the simultaneous and sensitive detection of multiple foodborne pathogens is problematic in microbial detection.Molecular biology and immunohistochemical analysis techniques have made some attempts in this detection field.Still,due to the limitations of primer design and antibodies,the effects of these two techniques in practical applications are not very satisfactory.Surface-enhanced Raman spectroscopy(SERS)technology has gained essential applications in the simultaneous detection of multiple pathogens due to its rapid,non-destructive,high-resolution,no interference from water,and in-situ detection.This review systematically summarizes the application strategies of SERS technology in the simultaneous detection of multiple pathogenic bacteria,including the application principle,application characteristics,and application effects.Firstly,a brief overview of the combination between the SERS substrate materials and the pathogens is introduced.Then the direct and indirect methods used in detecting multiple pathogens are presented separately.The direct method is simple and fast,and the spectral information of the pathogen itself is obtained directly through the signal amplification of the substrate material,which helps to study the information of the pathogen itself.It is widely used in multiple pathogenic bacteria discriminant,quantitative,and point-of-care testing(POCT).However,a large amount of spectral information,often needs to be used in conjunction with multivariate statistical analysis methods,imaging techniques,and microfluidic devices.Raman reporter molecules and recognition elements such as aptamers and antibodies are needed in the indirect method,converting the detection of pathogenic bacteria into the analysis of signal molecules.The significantly improves the sensitivity and specificity of the detection assay.Simultaneous analysis of multiple pathogenic bacteria can be achieved at the gene,protein,and cellular levels.Besides,a more comprehensive detection system that integrates bacteria’s separation,identification,and inactivation can be built by combining with other identification elements and functional molecules,which has essential prospects in analyzing multiple pathogens in actual samples such as clinical blood.Finally,the existing problems and following efforts of SERS technology are pointed out,which can be a reference for designing and applications of SERS technology in the rapid and sensitive detection of multiple pathogenic bacteria.