Herein, an autocatalytic strand displacement amplification (ASDA) strategy was proposed for the first time, which was further ingeniously coupled with hybridization chain reaction (HCR) event for the isothermal, label-free and multiple amplification toward nucleic acid detection. During the ASDA module, the target recognition opens the immobilized hairpin probe (IP) and initiates the annealing of the auxiliary DNA strand (AS) with the opened IP for the successive polymerization and nicking reaction in the presence of DNA polymerase and nicking endonuclease. This induces the target recycling and generation of a large amount of intermediate DNA sequences, which can be used as target analogy to execute the autocatalytic strand displacement amplification. Simultaneously, the introduced AS strand can propagate the HCR between two hairpins (H1 and H2) to form a linear DNA concatamer with cytosine (C)-rich loop region, which can facilitate the in-situ synthesis of silver nanoclusters (AgNCs) as electrochemical tags for further amplification toward target responses. With current cascade ASDA and HCR strategy, the detection of target DNA could be achieved with a low detection limit of about 0.16 fM and a good selectivity. The developed biosensor also exhibits the distinct advantages of flexibility and simplicity in probe design and biosensor fabrication, and label-free electrochemical detection, thus opens a promising avenue for the detection of nucleic acid with low abundance in bioanalysis and clinical biomedicine.

在本文中，首次提出了自动催化链置换扩增（ASDA）策略，该策略进一步巧妙地与杂交链反应（HCR）事件结合，用于等温，无标记和多重扩增，朝着核酸检测的方向发展。在ASDA模块中，目标识别打开固定的发夹探针（IP），并用打开的IP启动辅助DNA链（AS）的退火，以在DNA聚合酶和切口内切核酸酶存在下进行连续聚合和切口反应。这诱导了靶的再循环和大量中间DNA序列的产生，其可用作靶类比以执行自催化链置换扩增。同时，引入的AS链可以在两个发夹（H1和H2）之间传播HCR，以形成具有富含胞嘧啶（C）的环状区域的线性DNA辅酶，这可以促进原位合成银纳米簇（AgNC）作为电化学标记，进一步扩大到目标反应。使用当前的级联ASDA和HCR策略，可以以约0.16 fM的低检测限和良好的选择性实现目标DNA的检测。先进的生物传感器在探针设计和生物传感器制造以及无标记电化学检测方面也显示出灵活性和简单性的独特优势，从而为生物分析和临床生物医学中低丰度核酸的检测开辟了一条有希望的途径。可以促进原位合成银纳米簇（AgNCs）作为电化学标签，进一步向靶标反应扩增。使用当前的级联ASDA和HCR策略，可以以约0.16 fM的低检测限和良好的选择性实现目标DNA的检测。先进的生物传感器在探针设计和生物传感器制造以及无标记电化学检测方面也显示出灵活性和简单性的独特优势，从而为生物分析和临床生物医学中低丰度核酸的检测开辟了一条有希望的途径。可以促进原位合成银纳米簇（AgNCs）作为电化学标签，进一步向靶标反应扩增。使用当前的级联ASDA和HCR策略，可以以约0.16 fM的低检测限和良好的选择性实现目标DNA的检测。先进的生物传感器在探针设计和生物传感器制造以及无标记电化学检测方面也显示出灵活性和简单性的独特优势，从而为生物分析和临床生物医学中低丰度核酸的检测开辟了一条有希望的途径。16 fM和良好的选择性。先进的生物传感器在探针设计和生物传感器制造以及无标记电化学检测方面也显示出灵活性和简单性的独特优势，从而为生物分析和临床生物医学中低丰度核酸的检测开辟了一条有希望的途径。16 fM和良好的选择性。先进的生物传感器在探针设计和生物传感器制造以及无标记电化学检测方面也显示出灵活性和简单性的独特优势，从而为生物分析和临床生物医学中低丰度核酸的检测开辟了一条有希望的途径。