Qiao Qinqin, Guo Xiaodong, Wen Fang, Chen Lu, Xu Qingbiao, Zheng Nan, Cheng Jianbo, Xue Xiuheng, Wang Jiaqi
Laboratory of Quality and Safety Risk Assessment for Dairy Products of Ministry of Agriculture and Rural Affairs, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing, China.
College of Information Engineering, Fuyang Normal University, Fuyang, China.
Front Chem. 2021 Mar 24;9:653869. doi: 10.3389/fchem.2021.653869. eCollection 2021.
Aflatoxin M (AFM), one of the most toxic mycotoxins, is a feed and food contaminant of global concern. In this study, we developed a fast and simple method for detection of AFM based on a structure-switching signaling aptamer. This aptasensor is based on the change in fluorescence signal due to formation of an AFM/aptamer complex. To generate the aptasensor, the specific aptamer was modified with FAM (carboxyfluorescein), and their complementary DNAs (cDNA) were modified with a carboxytetramethylrhodamine (TAMRA) quenching group. In the absence of AFM, the aptamers were hybridized with cDNA, resulting in quenching of the aptamer fluorescence due to the proximity of the aptamer's fluorophore to the quenching group on the cDNA. On the other hand, in the presence of AFM, a structural switch in the aptamer was induced by formation of an AFM/aptamer complex. Changes in the structure of the aptamer led to the release of the cDNA, causing the generation of a fluorescence signal. Thus, AFM concentrations could be quantitatively monitored based on the changes in fluorescences. Under optimized conditions, this assay exhibited a linear response to AFM in the range of 1-100 ng/mL and a limit of detection of 0.5 ng/mL was calculated. This proposed aptasensor was applied to milk samples spiked with a dilution series of AFM, yielding satisfactory recoveries from 93.4 to 101.3%. These results demonstrated that this detection technique could be useful for high-throughput and quantitative determination of mycotoxin levels in milk and dairy products.
黄曲霉毒素M（AFM）是毒性最强的霉菌毒素之一，是一种全球关注的饲料和食品污染物。在本研究中，我们基于结构转换信号适体开发了一种快速简便的AFM检测方法。这种适体传感器基于AFM/适体复合物形成导致的荧光信号变化。为了生成适体传感器，将特异性适体用羧基荧光素（FAM）修饰，其互补DNA（cDNA）用羧基四甲基罗丹明（TAMRA）淬灭基团修饰。在没有AFM的情况下，适体与cDNA杂交，由于适体荧光团靠近cDNA上的淬灭基团，导致适体荧光淬灭。另一方面，在有AFM的情况下，AFM/适体复合物的形成诱导适体发生结构转换。适体结构的变化导致cDNA释放，从而产生荧光信号。因此，可以根据荧光变化定量监测AFM浓度。在优化条件下，该测定法对AFM在1-100 ng/mL范围内呈现线性响应，计算出的检测限为0.5 ng/mL。将该提出的适体传感器应用于添加了一系列稀释AFM的牛奶样品，回收率在93.4%至101.3%之间，结果令人满意。这些结果表明，这种检测技术可用于高通量定量测定牛奶和乳制品中的霉菌毒素水平。
