基于氮掺杂多孔碳的核酸适配体传感器检测赭曲霉毒素A

    DETECTION OF OCHRATOXIN A BASED ON NITROGEN DOPED POROUS CARBON-APTAMER SENSOR

    • 摘要: 利用电化学方法简单快速、仪器易于小型化等优势,结合核酸适配体的强亲和力、高稳定性等特点将其作为特异性识别元件固定在金电极(AuE)表面,利用新型纳米材料氮掺杂多孔碳的高比表面积固载互补链DNA(cDNA),通过杂交制备氮掺杂多孔碳-cDNA/核酸适配体/AuE传感器,以亚甲基蓝(MB)为电化学信号探针,进行OTA的检测研究。对制备传感器的主要参数进行优化,得到最佳试验条件:核酸适配体的孵育时间为2 h、氮掺杂多孔碳-cDNA浓度为5 μmol/L、核酸适配体与氮掺杂多孔碳-cDNA的杂交时间为1.5 h。该传感器用于OTA检测的结果表明,当加入OTA 5.0×10-6 μg/mL后,传感器的电流降低1.1 μA,这是因为OTA的存在使得核酸适配体构象发生变化形成OTA-核酸适配体复合物,导致氮掺杂多孔碳-cDNA从电极表面脱落,能与G碱基特异性结合的MB随之减少,从而使传感器表面MB峰值电流降低。OTA质量浓度在1.0×10-7~5.0×10-5μg/mL范围内具有良好的线性关系。研究表明,所制备的传感器具有良好的重现性、重复性和可接受的稳定性。利用加标法对大米样品中OTA进行检测,平均回收率为102%。

       

      Abstract: As the specific identification element,the aptamer of ochratoxin A was immobilized on the surface of gold electrode (AuE) based on its strong affinity and high stability.Using nitrogen doped porous carbon with a large specific surface area to carry more complementary strand DNA (cDNA),the hybridization was carried out on the AuE to obtain the nitrogen doped porous carbon-DNA/aptamer/AuE electrochemical sensor.The detection of OTA was studied by using methylene blue as the electrochemical signal probe.The main experimental parameters were optimized in the preparation process of the aptamer sensor.The optimal experimental conditions were as follows:the incubation time of the aptamer was 2 h,the concentration of the carbon aerogels-cDNA was 5 μmol/L,and the hybridization time between the aptamer and nitrogen doped porous carbon-cDNA was 1.5 h.The electric current was reduced by 1.1 μA when incubating with 5.0×10-6μg/mL OTA,and this signal change can be explained as follows:in the absence of OTA,MB can intercalate into single-stranded cDNA through the guanine bases and double-stranded DNA,and produce a strong current signal.In the presence of OTA,the binding of OTA and aptamer is considerably greater than that of cDNA and aptamer,which results in the release of MB from the electrode surface and reduces current signal.The linear equation was obtained in the OTA concentration range fnrm 1.0×10-7to 5.0×10-5μg/mL.The sensor had good reproducibility and repeatability,and acceptable stability.The rice samples were tested by the spiking method,and the average recovery was 102%.

       

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