An Aptamer-Array-Based Sample-to-Answer Biosensor for Ochratoxin A Detection via Fluorescence Resonance Energy Transfer

Yu Wang,Min Fang,Gongjun Tan,Tianfu Wu,Yaxi Li,Shu Zhang,Zhenfei Peng,Min Xiao,Bowen Yang,Wenlian Wang,Yongning Li,Chenling Tang

doi:10.3390/chemosensors9110309

Abstract

Food toxins are a hidden threat that can cause cancer and tremendously impact human health. Therefore, the detection of food toxins in a timely manner with high sensitivity is of paramount importance for public health and food safety. However, the current detection methods are relatively time-consuming and not practical for field tests. In the present work, we developed a novel aptamer-chip-based sample-to-answer biosensor (ACSB) for ochratoxin A (OTA) detection via fluorescence resonance energy transfer (FRET). In this system, a cyanine 3 (Cy3)-labeled OTA-specific biotinylated aptamer was immobilized on an epoxy-coated chip via streptavidin-biotin binding. A complementary DNA strand to OTA aptamer at the 3′-end was labeled with a black hole quencher 2 (BHQ2) to quench Cy3 fluorescence when in proximity. In the presence of OTA, the Cy3-labeled OTA aptamer bound specifically to OTA and led to the physical separation of Cy3 and BHQ2, which resulted in an increase of fluorescence signal. The limit of detection (LOD) of this ACSB for OTA was 0.005 ng/mL with a linearity range of 0.01–10 ng/mL. The cross-reactivity of ACSB against other mycotoxins, ochratoxin B (OTB), aflatoxin B1 (AFB1), zearalenone (ZEA), or deoxynilvalenol (DON), was less than 0.01%. In addition, this system could accurately detect OTA in rice samples spiked with OTA, and the mean recovery rate of the spiked-in OTA reached 91%, with a coefficient of variation (CV) of 8.57–9.89%. Collectively, the ACSB may represent a rapid, accurate, and easy-to-use platform for OTA detection with high sensitivity and specificity.

Highlights

Introduction published maps and institutional affilMycotoxins are secondary metabolites of fungi and can cause severe health problems [1]
Ochratoxin A (OTA) is a mycotoxin produced by several fungal species including Aspergillus ochraceus, A. carbonarius, A. niger and Penicillium verrucosum [2]
The mean fluorescence intensity was lower than 2000 a.u. when ochratoxin B (OTB) was at 10 ng/mL or lower, which was far below the mean fluorescence intensity of 40,000 a.u. when OTA was at 0.1~10 ng/mL. These results indicate that there was barely any cross-reactivity of the OTA sensor in detecting other mycotoxins including OTB

Summary

Introduction

Mycotoxins are secondary metabolites of fungi and can cause severe health problems [1]. There are growing concerns of food safety issues due to the contamination of mycotoxins. Ochratoxin A (OTA) is a mycotoxin produced by several fungal species including Aspergillus ochraceus, A. carbonarius, A. niger and Penicillium verrucosum [2]. OTA occurs in a wide variety of commonly contaminated agricultural commodities, such as grains, nuts, spices, coffee beans, olives, grapes, dried fruits, wine, and meat products [3,4]. OTA is notorious for its teratogenicity, embryotoxicity, genotoxicity, neurotoxicity, carcinogenicity, nephrotoxicity, and immunosuppressive effects [5]. OTA exposure could lead to adverse health effects in different populations, such as Balkan endemic nephropathy, chronic interstitial nephropathy, and other renal diseases [2].

Results

Discussion

Conclusion