Department of Laboratory Medicine, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, 400038, China; Department of Laboratory Medicine, The Second Affiliated Hospital of Chongqing Medical University, Chongqing, 400010, PR China.
Department of Laboratory Medicine, The First Affiliated Hospital of Chongqing Medical University, Chongqing, 400016, China.
Talanta. 2024 Jan 1;266(Pt 2):125034. doi: 10.1016/j.talanta.2023.125034. Epub 2023 Aug 5.
Circulating tumor DNA (ctDNA) is a noninvasive biomarker for liquid biopsy with important clinical and biological information, but existing detection techniques are expensive, complex and quite time-consuming. Here, we report an ultrarapid, sensitive and simple method, which we term Thermus thermophilus argonaute-coupling exponential amplification reaction (TtAgo-CEAR), that selectively amplifies mutated ctDNA. Aiming at seven Kirsten rat sarcoma-2 virus (KRAS) point mutations, the present strategy allows for easy detection with attomolar sensitivity and single-nucleotide specificity within as little as 16 min without prior PCR amplification. We also demonstrate that TtAgo-coupling assay is easily adaptable to Terahertz spectroscopy-based and lateral-flow-based readout. We show that the detected ctDNA concentrations by mouse models can respond to the variations of disease burden in serum samples. It is envisioned that this TtAgo-CEAR approach has great potential for rapid diagnosis and monitoring of diverse malignant tumors.
循环肿瘤 DNA（ctDNA）是液体活检的一种非侵入性生物标志物，具有重要的临床和生物学信息，但现有的检测技术昂贵、复杂且耗时较长。在这里，我们报告了一种超快速、敏感和简单的方法，我们称之为嗜热链球菌 Argonaute 偶联指数扩增反应（TtAgo-CEAR），该方法可以选择性地扩增突变的 ctDNA。针对七种 Kirsten 大鼠肉瘤病毒（KRAS）点突变，本策略可在无需 PCR 扩增的情况下，在 16 分钟内以皮摩尔灵敏度和单核苷酸特异性轻松检测。我们还证明，TtAgo 偶联测定法很容易适应太赫兹光谱和横向流动读取。我们表明，通过小鼠模型检测到的 ctDNA 浓度可以响应血清样本中疾病负担的变化。可以预见，这种 TtAgo-CEAR 方法具有快速诊断和监测多种恶性肿瘤的巨大潜力。
