Liu Jianghua, Chen Jiahui, Wu Di, Huang Mingquan, Chen Jian, Pan Ruiyuan, Wu Yongning, Li Guoliang
School of Food and Biological Engineering, Shaanxi University of Science and Technology, Xi'an 710021, China.
Institute for Global Food Security, School of Biological Sciences, Queen's University Belfast, 19 Chlorine Gardens, Belfast BT9 5DL, United Kingdom.
Anal Chem. 2021 Jul 27;93(29):10167-10174. doi: 10.1021/acs.analchem.1c01163. Epub 2021 Jul 19.
Surface-enhanced Raman scattering (SERS) has been recognized as a powerful tool for biosensors due to the ultrahigh sensitivity and unique fingerprint information. However, there are some limitations in trace target nucleic acid detection for the restricted signal-transducing and amplification strategies. Inspired by CRISPR/Cas12a with specific target DNA-activated collateral single-strand DNA (ssDNA) cleavage activity and liposome with signal molecule-loading properties, we first proposed a sensitive SERS-based on-site nucleic acid detection strategy mediated by CRISPR/Cas12a with -cleavage activity on ssDNA linkers utilized to capture liposomes. Liposomes loading two kinds of signal molecules, 4-nitrothiophenol (4-NTP) and cysteine, could achieve the dual-mode detection of target DNA with SERS and naked eye, respectively. The promptly amplified signals were initiated by the triggered breakdown of signal molecule-loaded liposomes. Emancipated 4-NTP, a biological-silent Raman reporter, would achieve highly selective and sensitive SERS measurement. Released cysteine induced the aggregation of plasmonic gold nanoparticles, leading to an obvious red to blue colorimetric shift to realize portable naked-eye detection. With this strategy, target nucleic acid concentration was dexterously converted into SERS and visualization signals and could be detected as low as 100 aM and 10 pM, respectively. The approach was also successfully applied to determine meat adulteration, achieving the detection of a low adulteration ratio in the complicated food matrix. We anticipate that this strategy will not only be regarded as a universal platform for the on-site detection of food authenticity but also broaden SERS application for the accurate determination of diverse biomarkers.
表面增强拉曼散射（SERS）因其超高灵敏度和独特的指纹信息，已被公认为是一种用于生物传感器的强大工具。然而，由于信号转导和放大策略受限，痕量目标核酸检测存在一些局限性。受具有特异性靶DNA激活的附带单链DNA（ssDNA）切割活性的CRISPR/Cas12a和具有信号分子负载特性的脂质体的启发，我们首次提出了一种基于SERS的灵敏的现场核酸检测策略，该策略由对用于捕获脂质体的ssDNA接头具有切割活性的CRISPR/Cas12a介导。负载4-硝基硫酚（4-NTP）和半胱氨酸这两种信号分子的脂质体，可分别通过SERS和肉眼实现对靶DNA的双模式检测。信号分子负载的脂质体的触发分解引发了迅速放大的信号。释放出的4-NTP，一种生物沉默的拉曼报告分子，将实现高选择性和灵敏的SERS测量。释放出的半胱氨酸诱导等离子体金纳米颗粒聚集，导致明显的从红色到蓝色的比色变化，以实现便携式肉眼检测。通过这种策略，靶核酸浓度被巧妙地转化为SERS和可视化信号，检测下限分别可达100 aM和10 pM。该方法还成功应用于测定肉类掺假，实现了在复杂食品基质中低掺假率的检测。我们预计，该策略不仅将被视为食品真实性现场检测的通用平台，还将拓宽SERS在准确测定多种生物标志物方面的应用。
