Parallel Electrophoretic Analysis of Segmented Samples On Chip for High-Throughput Determination of Enzyme Activities
Parallel Electrophoretic Analysis of Segmented Samples On Chip for High-Throughput Determination of Enzyme Activities
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DOI：
10.1021/ac101755y
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发表时间：
2010-11-15
影响因子：
7.4
通讯作者：
Kennedy, Robert T.
中科院分区：
文献类型：
--
作者：
Pei, Jian;Nie, Jing;Kennedy, Robert T.
毛细管电泳法结合了快速的分离速度和并行操作，在微细加工结构上取得了令人印象深刻的样品吞吐量。进一步提高产量的一个障碍是缺乏以与分析速度匹配的速度加载和注入单个样品的方法。为了解决这个问题，我们开发了一种微流控装置，其中以被氟碳油分割的纳升体积塞子的形式存储的样品被顺序地引入到三个电泳道的阵列中。由图案化表面化学和几何限制组成的微流控界面被用来从每个分段的流动通道中提取样品并转移到相应的电泳通道进行分离。荧光检测是通过使用配备电荷耦合设备的荧光显微镜对芯片成像来实现的。对该系统的表征表明，进样量受样品塞体积、进样时的流速和施加在电泳道上的电压的控制。对该系统进行了GTP酶检测。酶产物峰面积比与内标峰面积比的相对标准偏差为6%。峰间交叉污染为7%。在10分钟内实现了120个样品的处理。该系统的进一步开发可能会应用于高通量应用，如药物筛选。
Capillary electrophoresis (CE) on microfabricated structures has achieved impressive sample throughput by combining fast separation speed and parallel operations. One obstacle to further increasing throughput has been lack of methods for loading and injecting individual samples at a rate that matches analysis speed. To address this issue, we have developed a microfluidic device in which samples stored as nanoliter volume plugs segmented by a fluorocarbon oil are introduced sequentially to an array of three electrophoresis channels. A microfluidic interface consisting of patterned surface chemistry and geometric restriction was used to extract samples from each segmented flow channel and transfer to the respective electrophoresis channel for separation. Fluorescence detection was achieved by imaging the chip using a fluorescence microscope equipped with a charge-coupled device. Characterization of the system shows that injection volume is controlled by sample plug volume, flow rate during introduction, and voltage applied to the electrophoresis channel. The system was tested for a GTPase assay. Peak area ratios of enzyme product and internal standard had 6% relative standard deviations. Cross-contamination between peaks was 7%. Throughput of 120 samples in 10 min was achieved. Further development of the system may allow application to highthroughput applications such as drug screening.