Lab in a Tube: Ultrasensitive Detection of MicroRNAs at the Single-Cell Level and in Breast Cancer Patients Using Quadratic Isothermal Amplification
Lab in a Tube: Ultrasensitive Detection of MicroRNAs at the Single-Cell Level and in Breast Cancer Patients Using Quadratic Isothermal Amplification
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管中实验室：使用二次等温扩增在单细胞水平和乳腺癌患者中超灵敏检测 MicroRNA
DOI：
10.1021/ja311313b
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发表时间：
2013-03-27
影响因子：
15
通讯作者：
Xia, Fan
中科院分区：
文献类型：
--
作者：
Duan, Ruixue;Zuo, Xiaolei;Xia, Fan
通过合理设计具有高序列特异性的功能分子探针，利用灵敏的等温扩增，操作简单，我们开发了一种一锅发夹介导的二次酶扩增策略，用于检测microRNA （miRNA）。我们的方法对miR-21具有超高的灵敏度，在37℃下检测限为10 fM，在4℃下检测限为1 aM，对应于15 μ L样品中的9股miR-21，并且能够区分miRNA家族成员。更重要的是，当应用于MCF-7和PC3细胞系的粗提取，甚至是乳腺导管内癌和浸润性导管癌的患者组织时，该方法也具有敏感性和选择性。
Through rational design of a functional molecular probe with high sequence specificity that takes advantage of sensitive isothermal amplification with simple operation, we developed a one-pot hairpin-mediated quadratic enzymatic amplification strategy for microRNA (miRNA) detection. Our method exhibits ultrahigh sensitivity toward miR-21 with detection limits of 10 fM at 37 degrees C and 1 aM at 4 degrees C, which corresponds to nine strands of miR-21 in a 15 mu L sample, and it is capable of distinguishing among miRNA family members. More importantly, the proposed approach is also sensitive and selective when applied to crude extractions from MCF-7 and PC3 cell lines and even patient tissues from intraductal carcinoma and invasive ductal carcinoma of the breast.