Rapid Isolation and Detection of Exosomes and Associated Biomarkers from Plasma
Rapid Isolation and Detection of Exosomes and Associated Biomarkers from Plasma
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DOI：
10.1021/acsnano.7b00549
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发表时间：
2017-07-01
期刊：
影响因子：
17.1
通讯作者：
Heller, Michael J.
中科院分区：
文献类型：
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作者：
Ibsen, Stuart D.;Wright, Jennifer;Heller, Michael J.
在血液循环中发现的外切体是重要的癌症相关RNA和蛋白质生物标记物的主要来源，有望导致早期发现、液体活检和护理点诊断应用。不幸的是，由于它们的小尺寸(50-150 nm)和低密度，外切体很难从血浆中分离出来。目前的隔离方法是耗时的多步骤程序，不太可能转化为诊断应用。为了解决这个问题，我们展示了交流电动(ACE)微阵列芯片设备从未稀释的人类血浆样本中快速分离和回收胶质母细胞瘤外切体的能力。该装置需要一个小的血浆样本(30-50亩L)，并能够在15分钟内将外切体集中到血管紧张素转换酶微电极周围的高场区。简单的缓冲液洗涤去除了大量的血浆物质，使外切体集中在微电极上。整个分离过程和芯片上的荧光分析在不到30分钟的时间内完成，这使得随后的芯片上免疫荧光检测外体蛋白成为可能，并为RT-PCR分析提供了活的mRNA。这些结果表明，ACE设备能够简化分离和回收外切体的过程，显著减少处理步骤的数量和所需的时间。
Exosomes found in the circulation are a primary source of important cancer-related RNA and protein biomarkers that are expected to lead to early detection, liquid biopsy, and point-of-care diagnostic applications. Unfortunately, due to their small size (50-150 nm) and low density, exosomes are extremely difficult to isolate from plasma. Current isolation methods are time-consuming multistep procedures that are unlikely to translate into diagnostic applications. To address this issue, we demonstrate the ability of an alternating current electrokinetic (ACE) microarray chip device to rapidly isolate and recover glioblastoma exosomes from undiluted human plasma samples. The ACE device requires a small plasma sample (30-50 mu L) and is able to concentrate the exosomes into high-field regions around the ACE microelectrodes within 15 min. A simple buffer wash removes bulk plasma materials, leaving the exosomes concentrated on the microelectrodes. The entire isolation process and on-chip fluorescence analysis is completed in less than 30 min which enables subsequent on-chip immunofluorescence detection of exosomal proteins, and provides viable mRNA for RT-PCR analysis. These results demonstrate the ability of the ACE device to streamline the process for isolation and recovery of exosomes, significantly reducing the number of processing steps and time required.