Yang Ruiqi, Zhao Liping, Fang Mengqi, Kong Weijun, Luan Yunxia
Institute of Quality Standard and Testing Technology of BAAFS, Beijing 100097, China; Haiyuan College, School of Pharmaceutical Sciences, Kunming Medical University, Kunming, 650500, PR China; State Key Laboratory of Southwestern Chinese Medicine Resources, Pharmacy College, Chengdu University of Traditional Chinese Medicine, Chengdu 611137, China.
Institute of Quality Standard and Testing Technology of BAAFS, Beijing 100097, China.
Talanta. 2025 May 1;286:127496. doi: 10.1016/j.talanta.2024.127496. Epub 2025 Jan 3.
Alternariol (AOH) has attracted much attention as an emerging toxin in edible herbs that can pose potential carcinogenic risks to human. However, the rapid detection of AOH to ensure food safety remains a challenge. Here, a CRISPR-Cas12a-mediated aptamer-based sensor (aptasensor) was proposed for the sensitive quantification of AOH by using a personal glucose meter. First, fluorescent probes were used to validate crRNA sequences that could activate Cas12a trans-cleavage activity. Owing to the high affinity of aptamer, different concentrations of AOH were able to release different levels of Cas12a cleavage activity to cut magnetic bead-modified invertase probes. The free invertase was collected to catalyze hydrolysis of sucrose to glucose. After optimizing a series of key parameters, the constructed aptasensor with dual signal amplification of Cas12a and invertase allowed for highly-sensitive AOH detection in a linear range of 0.45-3000 ng/mL with a detection limit of 0.048 ng/mL. In addition, this aptasensor was successfully applied in edible herb samples with satisfactory recovery rates of 92.19-102.40 %. This CRISPR-Cas12a-mediated aptasensing platform provides a simple and powerful alternative for rapid quantitative detection of AOH, which is of great significance for food safety.
交链孢酚（AOH）作为食用草药中一种新兴的毒素，已引起广泛关注，它可能对人类构成潜在致癌风险。然而，快速检测AOH以确保食品安全仍然是一项挑战。在此，提出了一种基于CRISPR-Cas12a的适体传感器（aptasensor），通过使用个人血糖仪对AOH进行灵敏定量。首先，使用荧光探针验证可激活Cas12a反式切割活性的crRNA序列。由于适体具有高亲和力，不同浓度的AOH能够释放不同水平的Cas12a切割活性，以切割磁珠修饰的转化酶探针。收集游离的转化酶以催化蔗糖水解为葡萄糖。在优化一系列关键参数后，构建的具有Cas12a和转化酶双重信号放大功能的适体传感器能够在0.45-3000 ng/mL的线性范围内对AOH进行高灵敏度检测，检测限为0.048 ng/mL。此外，该适体传感器成功应用于食用草药样品，回收率令人满意，为92.19-102.40%。这种基于CRISPR-Cas12a的适体传感平台为AOH的快速定量检测提供了一种简单而强大的替代方法，对食品安全具有重要意义。
