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Fluorometric aptamer based assay for ochratoxin A based on the use of exonuclease III

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Abstract

This study describes an aptamer based assay for the mycotoxin ochratoxin A (OTA). The method is based on the use of an OTA-specific aptamer, exonuclease (Exo) III, SYBR Gold as a fluorescent probe, and a complementary strand that specifically combines with the aptamer. In the presence of OTA, the aptamer and OTA hybridize, thereby resulting in the formation of ssDNA, which is not digested by Exo III. Intense fluorescence is observed after addition of SYBR Gold (best measured at excitation/emission wavelengths of 495/540 nm). Fluorescence increases linearly with the log of the OTA concentration in the range from 8 to 1000 ng·mL−1. The detection limit is 4.7 ng·mL−1. The assay was applied to the determination of OTA in diluted [2%(v/v)] red wine, and recoveries and RSDs ranged between 93.5% and 113.8%, and between 3.2% and 5.7%, respectively.

In the presence of ochratoxin A (OTA), specific combinations of aptamer and OTA may occur and result in DNA double strands being untied, which avoids being digested by Exo III. Intense fluorescence is observed after SYBR Gold addition.

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Acknowledgements

This work was supported by the National Natural Science Foundation of China (Nos. 31460423 and 31360384), the department of Sciences & Technology of Jilin Province (20160520047JH) and the department of education of Jilin Province (2016252).

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Author notes

  1. Renjie Liu, Hua Wu and Lei Lv contributed equally to this work.

Authors and Affiliations

  1. Institute of food science and engineering, Jilin agricultural University, Changchun, 130118, China

    Renjie Liu & Hua Wu

  2. College of agriculture, Yanbian university, Yanji, 133002, China

    Hua Wu, Lei Lv, Chengbi Cui, Jin Feng & Zhijun Guo

  3. School of Electrical Engineering and Intelligentization, Dongguan University of Technology, Dongguan, 523808, China

    Xiaojiao Kang

Authors
  1. Renjie Liu
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  2. Hua Wu
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  3. Lei Lv
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  4. Xiaojiao Kang
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  5. Chengbi Cui
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  6. Jin Feng
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Correspondence to Zhijun Guo.

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Liu, R., Wu, H., Lv, L. et al. Fluorometric aptamer based assay for ochratoxin A based on the use of exonuclease III. Microchim Acta 185, 254 (2018). https://doi.org/10.1007/s00604-018-2786-6

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