CRISPR/Cas12a triggered SERS and naked eye dual-mode biosensor for ultrasensitive and on-site detection of nucleic acid via cascade signal amplification
CRISPR/Cas12a triggered SERS and naked eye dual-mode biosensor for ultrasensitive and on-site detection of nucleic acid via cascade signal amplification
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DOI：
10.1016/j.snb.2023.135249
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发表时间：
2024-01
期刊：
影响因子：
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通讯作者：
Jianghua Liu;Jiahui Chen;Shijie Jia;Yu Wang;Di Wu;Yongning Wu;Guoliang Li
中科院分区：
文献类型：
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作者：
Jianghua Liu;Jiahui Chen;Shijie Jia;Yu Wang;Di Wu;Yongning Wu;Guoliang Li
核酸的高灵敏和现场检测一直是分析化学领域的一个关键问题。基于表面增强拉曼散射(SERS)的生物传感器在核酸检测中显示出巨大的潜力，但由于缺乏合适的信号识别、转换和放大技术，其在痕量核酸筛选中的应用受到限制。受CRISPR/Cas12a的特异性识别和通过级联信号放大提高灵敏度的启发，我们创新性地提出了一种CRISPR/Cas12a触发的SERS和肉眼双模生物传感器，用于超灵敏和现场检测核酸的级联信号放大。在靶DNA识别后，激活的CRISPR/Cas12a不分青红皂白地切割底物ssDNA，导致脚趾介导的DNA链置换反应(TSDR)失败，并引发杂交链式反应(HCR)组装大量G-四链/氯化血红素DNAzyme(GQH DNAzyme)进行级联信号放大。生成的GQH DNAzyme催化l-半胱氨酸氧化为半胱氨酸，扰乱了4-NTP@AuNPs的聚集，导致了显著的拉曼信号变化。另一方面，GQH脱氧核酶催化2，2′-azino-di-(3-ethylbenzthiazoline-6-sulfonic酸(ABTS)的氧化，导致明显的颜色变化，实现便携式肉眼检测。通过这一策略，将目标核酸浓度巧妙地转换为灵敏的拉曼信号和便携式可视化信号，检测限分别低至34.9am和1pm。将该生物传感器成功应用于肉类掺假检测，对复杂食品基质中微量核酸的现场检测具有良好的选择性、灵敏度和适用性。
Highly sensitive and on-site detection of nucleic acids has always been a critical issue in the field of analytical chemistry. Surface-enhanced Raman scattering (SERS)-based biosensing exhibits huge potential in nucleic acid detection, while the applicability is restricted in trace nucleic acid screening due to the lack of appropriate signal recognition, transducing and amplification technologies. Inspired by the specific recognition of CRISPR/Cas12a and the improved sensitivity through cascade signal amplification, we innovatively proposed a CRISPR/Cas12a triggered SERS and naked eye dual-mode biosensor for ultrasensitive and on-site detection of nucleic acid via cascade signal amplification. Upon the target DNA recognition, the activated CRISPR/Cas12a indiscriminately cleaved substrate ssDNA, leading to the failure of toehold-mediated DNA-strand displacement reaction (TSDR), and triggering hybridization chain reaction (HCR) to assemble numerous G-quadruplex/hemin DNAzyme (GQH DNAzyme) for cascade signal amplification. The generated GQH DNAzyme catalyzed the oxidation ofl-cysteine to cystine, perturbing the aggregation of 4-NTP@AuNPs, resulting in significant Raman signal change. On the other hand, GQH DNAzyme catalyzed the oxidation of 2,2′-azino-di-(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), leading to obvious color change to realize portable naked-eye detection. Through this strategy, target nucleic acid concentration was tactfully transformed into sensitive Raman and portable visualization signals, and the limit of detection were as low as 34.9 aM and 1 pM, respectively. Then, this biosensor was successfully applied to meat adulteration detection, which showed superb selectivity, sensitivity and applicability for on-site detection of trace nucleic acid in complicated food matrix.