Yuzuncu Yil University, Department of Analytical Chemistry, Faculty of Pharmacy, 65010 Van, Turkey; Department of Chemistry "Ugo Schiff", University of Florence, Via della Lastruccia 3, 50019, Sesto Fiorentino, Florence, Italy.
Department of Chemistry "Ugo Schiff", University of Florence, Via della Lastruccia 3, 50019, Sesto Fiorentino, Florence, Italy.
Bioelectrochemistry. 2021 Apr;138:107691. doi: 10.1016/j.bioelechem.2020.107691. Epub 2020 Nov 5.
This work proposes a voltammetric aptasensor to detect deoxynivalenol (DON) mycotoxin. The development steps of the aptasensor were partnered for the first time to a computational study to gain insights onto the molecular mechanisms involved into the interaction between a thiol-tethered DNA aptamer (80mer-SH) and DON. The exploited docking study allowed to find the binding region of the oligonucleotide sequence and to determine DON preferred orientation. A biotinylated oligonucleotide sequence (20mer-BIO) complementary to the aptamer was chosen to carry out a competitive format. Graphite screen-printed electrodes (GSPEs) were electrochemically modified with polyaniline and gold nanoparticles (AuNPs@PANI) by means of cyclic voltammetry (CV) and worked as a scaffold for the immobilization of the DNA aptamer. Solutions containing increasing concentrations of DON and a fixed amount of 20mer-BIO were dropped onto the aptasensor surface: the resulting hybrids were labeled with an alkaline phosphatase (ALP) conjugate to hydrolyze 1-naphthyl phosphate (1-NPP) substrate into 1-naphthol product, detected by differential pulse voltammetry (DPV). According to its competitive format, the aptasensor response was signal-off in the range 5.0-30.0 ng·mL DON. A detection limit of 3.2 ng·mL was achieved within a 1-hour detection time. Preliminary experiments on maize flour samples spiked with DON yielded good recovery values.
本工作提出了一种用于检测脱氧雪腐镰刀菌烯醇（DON）真菌毒素的伏安适配体传感器。该适配体传感器的开发步骤首次与计算研究相结合，以深入了解与硫醇键合 DNA 适体（80mer-SH）和 DON 相互作用相关的分子机制。所利用的对接研究允许找到寡核苷酸序列的结合区域，并确定 DON 的优先取向。选择与适体互补的生物素化寡核苷酸序列（20mer-BIO）进行竞争性格式实验。通过循环伏安法（CV）将石墨丝网印刷电极（GSPE）电化学修饰为聚苯胺和金纳米粒子（AuNPs@PANI），并作为固定 DNA 适体的支架。将含有不同浓度 DON 和固定量 20mer-BIO 的溶液滴加到适配体传感器表面：所得杂交体用碱性磷酸酶（ALP）标记，以将 1-萘基磷酸酯（1-NPP）底物水解成 1-萘酚产物，通过差分脉冲伏安法（DPV）检测。根据其竞争性格式，适配体传感器在 5.0-30.0 ng·mL DON 范围内表现为信号关闭。在 1 小时的检测时间内，检测限达到 3.2 ng·mL。对 DON 污染的玉米粉样品进行的初步实验得到了良好的回收率值。
