Zhang Chi, Zhang Houchun, Wu Peng, Zhang Xinfeng, Liu Juewen
College of Materials and Chemistry & Chemical Engineering, Chengdu University of Technology, Chengdu, 610059, China.
Key Laboratory of Green Chemistry and Technology, Analytical & Testing Center, Sichuan University, 29 Wangjiang Road, Chengdu, 610064, China.
Biosens Bioelectron. 2020 Dec 1;169:112603. doi: 10.1016/j.bios.2020.112603. Epub 2020 Sep 9.
Peroxidase-like DNAzymes have been extensively used to replace horseradish peroxidase (HRP) for developing biosensors for signal amplification. However, the background activity from the cofactor (i.e., free hemin) has limited the sensitivity of such sensors. Herein, we aim to find an inhibitor for hemin to suppress the background signal, and a classic split DNAzyme-based sensor was used to detect a complementary DNA oligonucleotide. After screening a series of dyes, SYBR Green I (SG, one of the DNA stanning dyes) was selected for suppressing the background. Simply by adding 0.84 μM SG, the background from 50 nM hemin was suppressed over 30-fold. The suppression was caused by the interaction between SG and hemin. In the presence of the target DNA, the formed duplex region and G-quadruplex structure can better bind SG and hemin respectively, thus preventing the interaction between them and showing a high activity of the DNAzyme. The optimized sensor showed a detection limit of 3.8 pM for the target DNA (p53 gene). In addition, the backgrounds from chemiluminescence, colorimetric and fluorescence sensing modes can all be reduced by adding SG to the split DNAzyme system. The suppression of the background of peroxidase DNAzymes is a critical step towards practical use of related biosensors.
过氧化物酶样DNA酶已被广泛用于替代辣根过氧化物酶（HRP），以开发用于信号放大的生物传感器。然而，辅因子（即游离血红素）的背景活性限制了此类传感器的灵敏度。在此，我们旨在寻找一种血红素抑制剂来抑制背景信号，并使用一种基于经典分裂DNA酶的传感器来检测互补DNA寡核苷酸。在筛选了一系列染料后，选择了SYBR Green I（SG，一种DNA染色染料）来抑制背景。只需加入0.84 μM的SG，50 nM血红素的背景就被抑制了30倍以上。这种抑制是由SG与血红素之间的相互作用引起的。在目标DNA存在的情况下，形成的双链区域和G-四链体结构可以分别更好地结合SG和血红素，从而阻止它们之间的相互作用，并显示出DNA酶的高活性。优化后的传感器对目标DNA（p53基因）的检测限为3.8 pM。此外，通过向分裂DNA酶系统中添加SG，可以降低化学发光、比色和荧光传感模式的背景。抑制过氧化物酶DNA酶的背景是相关生物传感器实际应用的关键一步。
