An electrochemical clamp assay for direct, rapid analysis of circulating nucleic acids in serum
An electrochemical clamp assay for direct, rapid analysis of circulating nucleic acids in serum
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DOI：
10.1038/nchem.2270
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发表时间：
2015-07-01
期刊：
影响因子：
21.8
通讯作者：
Kelley, Shana O.
中科院分区：
文献类型：
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作者：
Das, Jagotamoy;Ivanov, Ivaylo;Kelley, Shana O.
无细胞核酸（cfNAs）在癌症患者的血液中含量很高，对其进行分析可以揭示肿瘤的突变谱，而无需对组织进行侵入性采样。然而，这需要区分来自健康细胞的核酸和来自肿瘤细胞的突变序列。在这里，我们报告了一种电化学钳法，可以直接检测患者血清中的突变序列。这是首次在不需要酶扩增的情况下成功检测cfNAs，酶扩增通常需要大量的样品处理并且容易受到干扰。这种新的基于芯片的检测方法使用一组寡核苷酸在15分钟内读出突变的存在，这些寡核苷酸将密切相关的序列隔离在溶液中，因此只允许突变序列与基于芯片的传感器结合。我们展示了出色的灵敏度和特异性，并表明钳法准确地检测出肺癌和黑色素瘤患者样本中的突变序列。
The analysis of cell-free nucleic acids (cfNAs), which are present at significant levels in the blood of cancer patients, can reveal the mutational spectrum of a tumour without the need for invasive sampling of the tissue. However, this requires differentiation between the nucleic acids that originate from healthy cells and the mutated sequences shed by tumour cells. Here we report an electrochemical clamp assay that directly detects mutated sequences in patient serum. This is the first successful detection of cfNAs without the need for enzymatic amplification, a step that normally requires extensive sample processing and is prone to interference. The new chip-based assay reads out the presence of mutations within 15 minutes using a collection of oligonucleotides that sequester closely related sequences in solution, and thus allow only the mutated sequence to bind to a chip-based sensor. We demonstrate excellent levels of sensitivity and specificity and show that the clamp assay accurately detects mutated sequences in a collection of samples taken from lung cancer and melanoma patients.