Qiao Bin, Xu Jiakun, Yin Wenhao, Xin Wanmeng, Ma Lixin, Qiao Jie, Liu Yi
Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Zhengzhou University, China.
Key Lab of Sustainable Development of Polar Fisheries, Ministry of Agriculture and Rural Affairs, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Lab for Marine Drugs and Byproducts of Pilot National Lab for Marine Science and Technology, China.
Biosens Bioelectron. 2021 Jul 1;183:113233. doi: 10.1016/j.bios.2021.113233. Epub 2021 Apr 7.
Herein, we report a method that combined "aptamer-locker" DNA with CRISPR/Cas12a-based biosensing for sensitive and rapid melamine analysis. Three strategies were harnessed for designing the DNA sensors that were well characterized by circular dichroism (CD) spectroscopy and isothermal titration calorimetry (ITC) in the absence and presence of melamine. The detection parameters were optimized to achieve good analytic performance. As a result, a limit of detection (LOD) as low as 38 nM was achieved, which is below the threshold (1.0 mg/kg) of allowable melamine in infant milk products. In addition, the sensors show high selectivity for melamine against other analogues such as cyanuric acid, ammeline and ammelide. Moreover, our method was effective for rapid melamine analysis in whole milk samples, with or without sample pretreatment, in less than 20 min. Adopting a commercially available portable fluorimeter, on-site analysis of melamine in milk was accomplished. The strategies demonstrated here can expand to detect other non-nucleic-acid targets by simply replacing the aptamers.
在此，我们报告了一种将“适体锁”DNA与基于CRISPR/Cas12a的生物传感相结合的方法，用于灵敏且快速地分析三聚氰胺。我们采用了三种策略来设计DNA传感器，在有无三聚氰胺的情况下，通过圆二色光谱（CD）和等温滴定量热法（ITC）对其进行了充分表征。对检测参数进行了优化，以实现良好的分析性能。结果，实现了低至38 nM的检测限（LOD），低于婴幼儿乳制品中三聚氰胺的允许阈值（1.0 mg/kg）。此外，该传感器对三聚氰胺具有高选择性，可区分其他类似物，如氰尿酸、三聚氰酸一酰胺和三聚氰酸二酰胺。而且，我们的方法可有效快速分析全脂牛奶样品中的三聚氰胺，无论有无样品预处理，均可在20分钟内完成。采用市售便携式荧光计，可实现牛奶中三聚氰胺的现场分析。此处展示的策略可通过简单更换适体扩展到检测其他非核酸靶标。
