State Key Laboratory of Food Science and Technology, School of Food Science, Jiangnan University, Wuxi, Jiangsu 214122, China.
J Agric Food Chem. 2012 Nov 7;60(44):10979-84. doi: 10.1021/jf3027233. Epub 2012 Oct 24.
A novel electrochemical DNA sensor was developed by using a stem-loop probe for peanut allergen Ara h 1 detection. The probe was modified with a thiol at its 5' end and a biotin at its 3' end. The biotin-tagged "molecular beacon"-like probe was attached to the surface of a gold electrode to form a stem-loop structure by self-assembly through facile gold-thiol affinity. 6-Mercaptohexanol (MCH) was used to cover the remnant bare region. The stem--loop probe was "closed" when the target was absent, and then the hybridization of the target induced the conformational change to "open", along with the biotin at its 3' end moved away from the electrode surface. The probe conformational change process was verified by circular dichroism (CD); meanwhile, electron-transfer efficiency changes between probe and electrode were proved by electrochemical impedance spectroscopy (EIS). The detection limit of this method was 0.35 fM with the linear response ranging from 10(-15) to 10(-10) M. Moreover, a complementary target could be discriminated from one-base mismatch and noncomplementarity. The proposed strategy has been successfully applied to detect Ara h 1 in the peanut DNA extracts of peanut milk beverage, and the concentration of it was 3.2 × 10(-13) mol/L.
一种新型电化学 DNA 传感器通过使用茎环探针用于检测花生过敏原 Ara h 1。该探针在其 5'端修饰有巯基，在其 3'端修饰有生物素。将生物素标记的“分子信标”样探针通过自组装通过简单的金-硫亲和力附着到金电极表面上以形成茎环结构。6-巯基己醇（MCH）用于覆盖剩余的裸露区域。当不存在靶标时，茎环探针“关闭”，然后靶标的杂交诱导构象变化为“打开”，同时其 3'端的生物素从电极表面移动。通过圆二色性（CD）验证了探针构象变化过程；同时，通过电化学阻抗谱（EIS）证明了探针和电极之间的电子转移效率变化。该方法的检测限为 0.35 fM，线性响应范围为 10(-15) 至 10(-10) M。此外，互补靶标可以与一个碱基错配和非互补性区分开来。该方法已成功应用于检测花生奶饮料中花生的 DNA 提取物中的 Ara h 1，其浓度为 3.2 × 10(-13) mol/L。
