Aptamer-Based Fluorescence Quenching Approach for Detection of Aflatoxin M1 in Milk

Qinqin Qiao; Xiaodong Guo; Fang Wen; Lu Chen; Qingbiao Xu; Nan Zheng; Jianbo Cheng; Xiuheng Xue; Jiaqi Wang

doi:10.3389/fchem.2021.653869

Aptamer-Based Fluorescence Quenching Approach for Detection of Aflatoxin M₁ in Milk

Front Chem. 2021 Mar 24:9:653869. doi: 10.3389/fchem.2021.653869. eCollection 2021.

Authors

Qinqin Qiao^{1

2

3}, Xiaodong Guo^{1

4

5}, Fang Wen^{1

5}, Lu Chen^{1

5}, Qingbiao Xu⁶, Nan Zheng^{1

5}, Jianbo Cheng³, Xiuheng Xue³, Jiaqi Wang^{1

5}

Affiliations

¹ 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.
² College of Information Engineering, Fuyang Normal University, Fuyang, China.
³ Anhui Agricultural University, Hefei, China.
⁴ School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, China.
⁵ Milk and Dairy Product Inspection Center of Ministry of Agriculture and Rural Affairs, Beijing, China.
⁶ College of Animal Sciences and Technology, Huazhong Agricultural University, Wuhan, China.

Abstract

Aflatoxin M₁ (AFM₁), 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 AFM₁ based on a structure-switching signaling aptamer. This aptasensor is based on the change in fluorescence signal due to formation of an AFM₁/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 AFM₁, 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 AFM₁, a structural switch in the aptamer was induced by formation of an AFM₁/aptamer complex. Changes in the structure of the aptamer led to the release of the cDNA, causing the generation of a fluorescence signal. Thus, AFM₁ concentrations could be quantitatively monitored based on the changes in fluorescences. Under optimized conditions, this assay exhibited a linear response to AFM₁ 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 AFM₁, 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.

Keywords: aflatoxin M1; aptasensor; fluorescence; food safety; quenching.