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首页> 外文期刊>Electroanalysis >Proximity Ligation Assay-induced Structure-switching Hairpin DNA toward Development of Electrochemical Immunosensor
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Proximity Ligation Assay-induced Structure-switching Hairpin DNA toward Development of Electrochemical Immunosensor

机译：邻近连接测定诱导结构转换发夹DNA向电化学免疫传感器的发展。

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A novel electrochemical immunosensor was designed for sensitive detection of mycotoxins (aflatoxin B-1, AFB(1), used in this case) by coupling proximity ligation assay-induced conformation switch of hairpin DNA with the antigen-antibody reaction. The assay was carried out by anti-AFB(1) antibody-conjugated DNA(1) (mAb-DNA(1)), AFB(1)-BSA-labeled DNA(2) (AFB(1)-DNA(2)) and hairpin DNA. Each hairpin included a stem of 6 base pairs and a 6 nucleotide (nt) loop with the labelled ferrocene tag at the 3 end, which was immobilized on the electrode via self-assembly of the terminal thiol moiety at the 5 end. The proximity ligation assay was carried out via the specific antigen-antibody reaction between mAb-DNA(1) and AFB(1)-DNA(2) to form an omega-like DNA junction. Thereafter, the junction hybridized with hairpin DNA to open the probe, thus resulting in the ferrocene tag far away from the electrode for the decreasing of the redox current. Upon target AFB(1) introduction, the competitive-type immunoassay was executed between the analyte and AFB(1)-DNA(2) for mAb-DNA(1). Under the optimal conditions, the electrochemical signal was indirectly proportional to target AFB(1) concentration, and allowed the detection of target AFB(1) at a concentration as low as 3.2pgmL(-1). Our strategy also exhibited high selectivity, good reproducibility and precision. Importantly, the accuracy of this methodology was validated for analysis of spiked or naturally contaminated peanut samples, giving results matched well with those obtained from commercialized available AFB(1) ELISA kit.
机译：一种新型的电化学免疫传感器被设计用于通过将邻近结扎法诱导的发夹DNA构象转换与抗原抗体反应偶联来灵敏检测霉菌毒素（在这种情况下使用的是黄曲霉毒素B-1，AFB（1））。通过抗AFB（1）抗体缀合的DNA（1）（mAb-DNA（1）），AFB（1）-BSA标记的DNA（2）（AFB（1）-DNA（2））进行测定）和发夹DNA。每个发夹包括6个碱基对的茎和在3末端带有标记的二茂铁标签的6个核苷酸（nt）环，其通过在5末端的末端硫醇部分的自组装而固定在电极上。通过mAb-DNA（1）和AFB（1）-DNA（2）之间的特异性抗原-抗体反应进行邻近连接测定，以形成类似ω的DNA连接。此后，该接头与发夹DNA杂交以打开探针，从而导致二茂铁标签远离电极，从而降低了氧化还原电流。在目标AFB（1）引入后，针对mAb-DNA（1）在分析物和AFB（1）-DNA（2）之间执行竞争型免疫分析。在最佳条件下，电化学信号与目标AFB（1）浓度成间接比例关系，并允许以低至3.2pgmL（-1）的浓度检测目标AFB（1）。我们的策略还表现出高选择性，良好的重现性和精度。重要的是，该方法的准确性已得到验证，可用于分析加标样品或自然污染的花生样品，其结果与从商业上可用的AFB（1）ELISA试剂盒获得的结果相匹配。

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