Zhou Qing-Yun, Ma Rong-Na, Hu Chao-Long, Sun Fei, Jia Li-Ping, Zhang Wei, Shang Lei, Xue Qing-Wang, Jia Wen-Li, Wang Huai-Sheng
School of Chemistry and Chemical Engineering, Liaocheng University, Liaocheng 252000, Shandong, P.R. China.
Oncology Department, Hospital of Traditional Chinese Medicine of Jinan City, Jinan 250000, Shandong, P.R. China.
Analyst. 2021 Apr 26;146(8):2705-2711. doi: 10.1039/d1an00204j.
A novel ratiometric electrochemical biosensing strategy based on T7 exonuclease (T7 Exo)-assisted homogenous target recycling coupling hairpin assembly triggered dual-signal output was proposed for the accurate and sensitive detection of microRNA-141 (miRNA-141). Concretely, in the presence of target miRNA, abundant signal transduction probes were released via the T7 Exo-assisted homogenous target recycling amplification, which could be captured by the specially designed ferrocene-labeled hairpin probe (Fc-H1) on -electrode interface and triggered the nonenzymatic catalytic hairpin assembly (Fc-H1 + MB-H2) to realize the cascade signal amplification and dual-signal output. Through such a conformational change process, the electrochemical signal of Fc (IFc) and MB (IMB) is proportionally and substantially decreased and increased. Therefore, the signal ratio of IMB/IFc can be employed to accurately reflect the true level of original miRNA. Benefiting from the efficient integration of the T7 Exo-assisted target recycle, nonenzymatic hairpin assembly and dual-signal output mode, the proposed sensor could realize the amplified detection of miRNA-141 effectively with a wide detection range from 1 fM to 100 pM, and a detection limit of 200 aM. Furthermore, it exhibits outstanding sequence specificity to discriminate mismatched RNA, acceptable reproducibility and feasibility for real sample. This strategy effectively integrated the advantages of multiple amplification and ratiometric output modes, which could provide an accurate and efficient method in biosensing and clinical diagnosis.
提出了一种基于T7核酸外切酶（T7 Exo）辅助的均相靶标循环耦合发夹组装触发双信号输出的新型比率电化学传感策略，用于准确、灵敏地检测微小RNA-141（miRNA-141）。具体而言，在靶标miRNA存在的情况下，通过T7 Exo辅助的均相靶标循环扩增释放大量信号转导探针，这些探针可被电极界面上专门设计的二茂铁标记发夹探针（Fc-H1）捕获，并触发非酶催化发夹组装（Fc-H1 + MB-H2）以实现级联信号放大和双信号输出。通过这样的构象变化过程，Fc（IFc）和亚甲基蓝（IMB）的电化学信号成比例且大幅降低和增加。因此，IMB/IFc的信号比可用于准确反映原始miRNA的真实水平。得益于T7 Exo辅助靶标循环、非酶发夹组装和双信号输出模式的有效整合，所提出的传感器能够在1 fM至100 pM的宽检测范围内有效地实现对miRNA-141的放大检测，检测限为200 aM。此外，它对错配RNA具有出色的序列特异性，具有可接受的重现性以及对实际样品检测的可行性。该策略有效整合了多种放大和比率输出模式的优势，可为生物传感和临床诊断提供一种准确、高效的方法。
