Exodisc for Rapid, Size-Selective, and Efficient Isolation and Analysis of Nanoscale Extracellular Vesicles from Biological Samples
Exodisc for Rapid, Size-Selective, and Efficient Isolation and Analysis of Nanoscale Extracellular Vesicles from Biological Samples
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DOI：
10.1021/acsnano.6b06131
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发表时间：
2017-02-01
期刊：
影响因子：
17.1
通讯作者：
Cho, Yoon-Kyoung
中科院分区：
文献类型：
--
作者：
Woo, Hyun-Kyung;Sunkara, Vijaya;Cho, Yoon-Kyoung
细胞外囊泡（EV）是细胞衍生的纳米级囊泡，其携带来自其来源细胞的核酸和蛋白质，并且显示出作为许多疾病（包括癌症）的生物标志物的巨大潜力。有效的分离和检测方法是利用其在临床环境中的使用和了解其生理功能的先决条件。在这里，我们提出了一种快速，无标记，高灵敏度的EV分离和定量方法，使用实验室上的一个光盘集成了两个纳米过滤器（Exodisc）。从原始生物样品开始，例如细胞培养上清液（CCS）或癌症患者尿液，使用桌面大小的离心微流体系统在30分钟内实现20-600 nm尺寸范围内的EV的全自动富集。使用纳米颗粒跟踪分析的定量测试证实，Exodisc能够从CCS中回收>95%的EV。此外，从EV中回收的mRNA的分析显示，与金标准超离心方法相比，Exodisc提供了>100倍的mRNA浓度。此外，使用从膀胱癌患者中分离的尿液EV进行的盘上酶联免疫吸附测定显示出高水平的CD 9和CD 81表达，这表明该方法可能在临床环境中潜在地用于测试用于癌症诊断的尿液EV生物标志物。
Extracellular vesicles (EVs) are cell-derived, nano scale vesicles that carry nucleic acids and proteins from their cells of origin and show great potential as biomarkers for many diseases, including cancer. Efficient isolation and detection methods are prerequisites for exploiting their use in clinical settings and understanding their physiological functions. Here, we presented a rapid, label-free, and highly sensitive method for EV isolation and quantification using a lab-on-a-disc integrated with two nanofilters (Exodisc). Starting from raw biological samples, such as cell-culture supernatant (CCS) or cancer patient urine, fully automated enrichment of EVs in the size range of 20-600 nm was achieved within 30 min using a tabletop-sized centrifugal microfluidic system. Quantitative tests using nanoparticle-tracking analysis confirmed that the Exodisc enabled >95% recovery of EVs from CCS. Additionally, analysis of mRNA retrieved from EVs revealed that the Exodisc provided >100-fold higher concentration of mRNA as compared with the gold-standard ultracentrifugation method. Furthermore, on-disc enzyme-linked immunosorbent assay using urinary EVs isolated from bladder cancer patients showed high levels of CD9 and CD81 expression, suggesting that this method may be potentially useful in clinical settings to test urinary EV-based biomarkers for cancer diagnostics.