State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210023, P.R. China.
Analyst. 2017 Sep 25;142(19):3740-3746. doi: 10.1039/c7an00413c.
A target-induced cyclic strategy for DNAzyme formation was proposed to achieve simple, sensitive and universal detection of protein biomarkers with convenient colorimetric or chemiluminescence imaging readout. In the assay, the target protein was recognized by a pair of DNA-labeled antibodies (Ab1-DNA1 and Ab2-DNA2) to form a proximate complex, which could hybridize with the conjugate DNA3/DNA4 to release the guanine-rich DNA4 and thus formed G-quadruplex/hemin horseradish peroxidase-mimicking DNAzyme. The process could be further recycled with Exonuclease III by cleaving DNA3 to free the proximate complex, resulting in the cyclic formation of DNAzyme. The G-quadruplex/hemin DNAzyme could catalyze the HO-mediated oxidation of 3,3,5,5-tetramethylbenzidine to produce the color change from colorless to blue or enhance the chemiluminescence of a luminol-HO system. Thus the signal could be read out with the naked eye, and by colorimetry and chemiluminescence imaging. Using a carcinoembryonic antigen as a model target, the proposed assay showed a detection range of 4 orders of magnitude along with detection limits of 170 and 16 pg mL for colorimetric and chemiluminescence imaging assays respectively. This assay had the advantages of easy operation, sensitive detection, target flexibility and diversified signal readout, providing a great opportunity for commercial application.
提出了一种基于靶诱导循环策略的 DNA zyme 形成方法，用于实现简单、灵敏且通用的蛋白质生物标志物检测，具有方便的比色或化学发光成像读出功能。在该测定法中，目标蛋白被一对 DNA 标记的抗体（Ab1-DNA1 和 Ab2-DNA2）识别，形成一个临近复合物，该复合物可以与结合 DNA3/DNA4 杂交，释放富含鸟嘌呤的 DNA4，从而形成 G-四链体/血红素辣根过氧化物酶模拟 DNAzyme。该过程可以通过 Exonuclease III 切割 DNA3 来进一步循环，从而释放出临近复合物，导致 DNAzyme 的循环形成。G-四链体/血红素 DNAzyme 可以催化 HO 介导的 3,3,5,5-四甲基联苯胺的氧化，产生从无色到蓝色的颜色变化，或增强鲁米诺-HO 体系的化学发光。因此，可以通过肉眼、比色法和化学发光成像来读取信号。使用癌胚抗原作为模型靶标，该测定法显示出 4 个数量级的检测范围，比色和化学发光成像测定法的检测限分别为 170 和 16 pg mL。该测定法具有操作简单、灵敏检测、靶标灵活性和多样化信号读出等优点，为商业应用提供了很好的机会。
