School of Pharmaceutical Sciences, Sun Yat-sen University, Guangzhou 510006, P. R. China.
Guangdong Second Provincial General Hospital, Guangzhou 510317, P. R. China.
Anal Chem. 2022 Jun 7;94(22):7918-7927. doi: 10.1021/acs.analchem.2c00723. Epub 2022 May 20.
The diagnosis of multidrug-resistant tuberculosis (MDR-TB) is crucial for the subsequent drug guidance to improve therapy and control the spread of this infectious disease. Herein, we developed a novel florescence biosensor for simultaneous detection of (Mtb) multidrug-resistant genes (rpoB531 for rifampicin and katG315 for isoniazid) by using our synthesized nanocobalt 5,10,15,20-tetra(4-pyridyl)-21,23-porphine (nanoCoTPyP) and double quantum dots (QDs). Several nanoCoTPyPs with different charges and morphology were successfully prepared via the surfactant-assisted method and their quenching ability and restoring efficiency for DNA detection were systematically analyzed. It was found that spherical nanoCoTPyP with positive charge exhibited excellent quenching effect and sensing performance for the two DNAs' detection due to its affinity differences towards single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA). ssDNA attached on QDs (QDs-ssDNA) was specifically hybridized with targets to form QDs-dsDNA, resulting in fluorescence recovery due to the disruption of the interactions between nanoCoTPyP and ssDNA. Two drug-resistant genes could be simultaneously quantified in a single run and relatively low limits of detection (LODs) were obtained (24 pM for T1 and 20 pM for T2). Furthermore, the accuracy and reliability of our method were verified by testing clinical samples. This simple and low-cost approach had great potential to be applied in clinical diagnosis of MDR-TB.
耐多药结核病（MDR-TB）的诊断对于后续的药物指导至关重要，有助于改善治疗效果并控制这种传染病的传播。在此，我们通过使用我们合成的纳米钴 5,10,15,20-四（4-吡啶基）-21,23-卟啉（nanoCoTPyP）和双量子点（QDs），开发了一种用于同时检测（Mtb）耐多药基因（rpoB531 用于利福平，katG315 用于异烟肼）的新型荧光生物传感器。通过表面活性剂辅助法成功制备了具有不同电荷和形态的几种 nanoCoTPyP，并系统地分析了它们对 DNA 检测的淬灭能力和恢复效率。结果发现，由于对单链 DNA（ssDNA）和双链 DNA（dsDNA）的亲和力差异，带正电荷的球形 nanoCoTPyP 对两种 DNA 的检测具有出色的淬灭效果和传感性能。附着在 QDs 上的 ssDNA（QDs-ssDNA）与靶标特异性杂交形成 QDs-dsDNA，由于 nanoCoTPyP 与 ssDNA 之间的相互作用被破坏，导致荧光恢复。两种耐药基因可以在单个运行中同时定量，并且获得了相对较低的检测限（T1 为 24 pM，T2 为 20 pM）。此外，通过测试临床样本验证了我们方法的准确性和可靠性。这种简单且低成本的方法具有在 MDR-TB 临床诊断中应用的巨大潜力。
