Simultaneous isolation and preconcentration of exosomes by ion concentration polarization.
Simultaneous isolation and preconcentration of exosomes by ion concentration polarization.
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DOI：
10.1002/elps.201700491
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发表时间：
2018-02-27
期刊：
影响因子：
2.9
通讯作者：
Chang HC
中科院分区：
文献类型：
--
作者：
Marczak S;Richards K;Ramshani Z;Smith E;Senapati S;Hill R;Go DB;Chang HC
外泌体携带microRNA生物标志物，在癌症患者中的丰度高于健康患者，并且因为它们存在于大多数生物流体中，包括血液和尿液，所以可以非侵入性地获得。分离外泌体的标准实验室技术昂贵、耗时、提供差的纯度，并且回收约25%的可用外泌体。我们提出了一种新的微流控技术，同时分离外泌体和预浓缩它们通过电泳使用高横向局部电场产生的离子耗尽离子选择性膜。我们使用压力驱动流将外泌体样品递送到微流体芯片，使得横向电场迫使它们离开交叉流并进入琼脂糖凝胶，该凝胶过滤掉不需要的细胞碎片，而离子选择性膜通过富集效应浓缩外泌体。我们从1X PBS缓冲液、细胞培养基和血清中有效地分离了外泌体。使用150 µL/hr至200 µL/hr的流速和100 V/cm的场强，我们一致地从缓冲液、细胞培养基和血清中捕获了60%至80%的外泌体，如荧光光谱和纳米颗粒跟踪分析所证实的。我们的微流控芯片保持这种回收率超过20分钟，10分钟的分离浓缩因子为15。
Exosomes carry microRNA biomarkers, occur in higher abundance in cancerous patients than in healthy ones, and because they are present in most biofluids, including blood and urine, can be obtained non-invasively. Standard laboratory techniques to isolate exosomes are expensive, time-consuming, provide poor purity, and recover on the order of 25% of the available exosomes. We present a new microfluidic technique to simultaneously isolate exosomes and preconcentrate them by electrophoresis using a high transverse local electric field generated by ion-depleting ion-selective membrane. We use pressure-driven flow to deliver an exosome sample to a microfluidic chip such that the transverse electric field forces them out of the cross flow and into an agarose gel which filters out unwanted cellular debris while the ion-selective membrane concentrates the exosomes through an enrichment effect. We efficiently isolated exosomes from 1X PBS buffer, cell culture media and blood serum. Using flow rates from 150 µL/hr to 200 µL/hr and field strengths of 100 V/cm, we consistently captured between 60% to 80% of exosomes from buffer, cell culture media, and blood serum as confirmed by both fluorescence spectroscopy and nanoparticle tracking analysis. Our microfluidic chip maintained this recovery rate for more than twenty minutes with a concentration factor of 15 for ten minutes of isolation.
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