Research Center of Analytical Instrumentation, Key Laboratory of Synthetic and Natural Functional Molecule Chemistry of Ministry of Education, College of Chemistry & Materials Science, Northwest University, Xi'an, 710069, Shaanxi, PR China; Research Center of Analytical Instrumentation, Key Laboratory of Bio-Resource and Eco-Environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu, 610065, Sichuan, PR China.
Research Center of Analytical Instrumentation, Key Laboratory of Bio-Resource and Eco-Environment of Ministry of Education, College of Life Sciences, Sichuan University, Chengdu, 610065, Sichuan, PR China.
Talanta. 2020 Apr 1;210:120638. doi: 10.1016/j.talanta.2019.120638. Epub 2019 Dec 12.
The cancer diagnosis with single level of biomarkers suffers from limitation of insufficient accuracy. Hence, developing sensitive, rapid and adaptative analytical strategies for double-level biomarkers are essential for improving the accuracy of clinical cancer diagnosis at early stage. Herein, a DNA biosensor was established based on the catalytic hairpin assembly-mediated Y-junction nicking enzyme assisted signal amplification (CHA-YNEASA) circuits, where the two circuits were concatenated by molecular beacon (MB). In absence of target, both the CHA and YNEASA circuits were effectively hindered because of MB's outstanding ability to control signal background. In presence of target, the initiated CHA circuits made enzyme recognition sequences in close proximity to the assisted sequences to open MB, leading to further trigger the YNEASA circuits. Due to the unique design of dual signal amplification strategies, CHA-YNEASA circuits significantly shorten the reaction time, and improve signal-to-background ratio as well as facilitate the analysis process. It was demonstrated that a high sensitivity with limit of detection (LOD) of 0.9 pM for p53 gene detection was obtained just within 23 min by the proposed DNA biosensor. Moreover, mismatched p53 gene at nucleic acid level was effectively discriminated and strong anti-interference capability was achieved. Noticeably, the DNA biosensor was adaptative for designing a cytosensor at cell level using hairpin DNA, containing MUC1 aptamer and initiation strand of CHA-YNEASA circuits, as switch based on modularity principle. The cytosensor is able to measure MUC1 positive breast cancer cells (MCF-7) with the LOD as low as 100 cells/mL. Excellent specificity for MUC1 negative cells, and good anti-interference capability in 10% fetal bovine serum (FBS) were observed by the cytosensor. Therefore, the proposed DNA biosensor is a sensitive, rapid, adaptative platform for detection of double-level biomarkers, offering novel strategy applied for clinical cancer diagnosis.
基于催化发夹组装介导的 Y 型分支核酸酶辅助信号放大（CHA-YNEASA）电路的 DNA 生物传感器的建立，其中两个电路通过分子信标（MB）串联。在不存在靶标时，由于 MB 出色的控制信号背景的能力，CHA 和 YNEASA 电路都有效地受到阻碍。在存在靶标的情况下，启动的 CHA 电路使酶识别序列与辅助序列紧密接近，从而打开 MB，进一步触发 YNEASA 电路。由于双信号放大策略的独特设计，CHA-YNEASA 电路显著缩短了反应时间，提高了信号与背景的比值，并促进了分析过程。实验证明，通过所提出的 DNA 生物传感器，仅在 23 分钟内即可实现 p53 基因检测的高灵敏度，检测限（LOD）为 0.9 pM。此外，在核酸水平上有效区分了错配的 p53 基因，并实现了强大的抗干扰能力。值得注意的是，该 DNA 生物传感器适用于使用发夹 DNA 设计细胞水平的细胞传感器，该发夹 DNA 包含 MUC1 适体和 CHA-YNEASA 电路的起始链，作为基于模块化原理的开关。该细胞传感器能够以低至 100 个细胞/mL 的检测限测量 MUC1 阳性乳腺癌细胞（MCF-7）。细胞传感器观察到对 MUC1 阴性细胞的优异特异性，以及在 10%胎牛血清（FBS）中的良好抗干扰能力。因此，所提出的 DNA 生物传感器是一种用于检测双水平生物标志物的灵敏、快速、自适应平台，为临床癌症诊断提供了新的策略。
