Quantification of Plasma miRNAs by Digital PCR for Cancer Diagnosis
Quantification of Plasma miRNAs by Digital PCR for Cancer Diagnosis
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DOI：
10.4137/bmimi.s13154
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发表时间：
2013-01-01
影响因子：
3.8
通讯作者：
Jiang, Feng
中科院分区：
文献类型：
--
作者：
Ma, Jie;Li, Ning;Jiang, Feng
通过定量聚合酶链反应（qPCR）分析血浆microRNA（miRNAs）为癌症诊断提供了潜在的方法。然而，用qPCR绝对定量低丰度血浆miRNA是具有挑战性的。数字PCR提供了一种独特的手段，用于评估血浆中低水平的核酸。本研究旨在评估数字PCR定量血浆miRNA的有效性以及该技术用于癌症诊断的潜在效用。我们使用数字PCR定量36例肺癌患者和38例对照者血浆microRNA-21- 5 p（miR-21- 5 p）和microRNA-335- 3 p（miR-335- 3 p）的拷贝数。数字PCR显示了高度的线性和定量相关性与miRNAs的动态范围从1到10,000拷贝/μ L的输入，具有高重现性。qPCR表现出100至1 × 107拷贝/μ L输入的动态范围。数字PCR检测miRNAs拷贝数的灵敏度高于qPCR。在血浆中，数字PCR可以检测miR-21- 5 p和miR-335- 3 p的拷贝数，而qPCR只能评估miR-21- 5 p。通过数字PCR定量血浆miRNA在区分肺癌患者和无癌受试者方面提供了71.8%的灵敏度和80.6%的特异性。
Analysis of plasma microRNAs (miRNAs) by quantitative polymerase chain reaction (qPCR) provides a potential approach for cancer diagnosis. However, absolutely quantifying low abundant plasma miRNAs is challenging with qPCR. Digital PCR offers a unique means for assessment of nucleic acids presenting at low levels in plasma. This study aimed to evaluate the efficacy of digital PCR for quantification of plasma miRNAs and the potential utility of this technique for cancer diagnosis. We used digital PCR to quantify the copy number of plasma microRNA-21-5p (miR-21-5p) and microRNA-335-3p (miR-335-3p) in 36 lung cancer patients and 38 controls. Digital PCR showed a high degree of linearity and quantitative correlation with miRNAs in a dynamic range from 1 to 10,000 copies/mu L of input, with high reproducibility. qPCR exhibited a dynamic range from 100 to 1X107 copies/mu L of input. Digital PCR had a higher sensitivity to detect copy number of the miRNAs compared with qPCR. In plasma, digital PCR could detect copy number of both miR-21-5p and miR-335-3p, whereas qPCR was only able to assess miR-21-5p. Quantification of the plasma miRNAs by digital PCR provided 71.8% sensitivity and 80.6% specificity in distinguishing lung cancer patients from cancer-free subjects.