High Content Cell Screening in a Microfluidic Device
High Content Cell Screening in a Microfluidic Device
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DOI：
10.1074/mcp.m800291-mcp200
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发表时间：
2009-03-01
影响因子：
7
通讯作者：
Levchenko, Andre
中科院分区：
文献类型：
--
作者：
Cheong, Raymond;Wang, Chiaochun Joanne;Levchenko, Andre
对细胞信号网络的全面系统级理解需要在单细胞水平上有效地分析多种信号物种，以响应各种刺激方案。在这里，我们描述了一种微流控装置，可以使用基于免疫荧光的读数对数千个单个细胞中的信号网络进行定量询问。该装置特别适用于以高通量、高含量的方式测量激酶、转录因子和/或靶基因的信号活性。我们演示了如何使用该设备来测量对一种或多种可溶性刺激和/或化学抑制剂的信号反应的详细时间过程，以及对多种刺激的复杂时间模式的反应。此外，我们还展示了如何利用该设备的吞吐量和分辨率来研究单细胞水平与群体水平上的信号传导差异（如果有的话）。特别是，我们表明，与过度表达p65-EGFP的细胞研究相比，单个细胞中的NF-kappa B活性动态不是异步的，而是类似于群体平均动态。中国生物医学工程学报（英文版），2009。
A comprehensive, systems level understanding of cell signaling networks requires methods to efficiently assay multiple signaling species, at the level of single cells, responding to a variety of stimulation protocols. Here we describe a microfluidic device that enables quantitative interrogation of signaling networks in thousands of individual cells using immunofluorescence-based readouts. The device is especially useful for measuring the signaling activity of kinases, transcription factors, and/or target genes in a high throughput, high content manner. We demonstrate how the device may be used to measure detailed time courses of signaling responses to one or more soluble stimuli and/or chemical inhibitors as well as responses to a complex temporal pattern of multiple stimuli. Furthermore we show how the throughput and resolution of the device may be exploited in investigating the differences, if any, of signaling at the level of a single cell versus at the level of the population. In particular, we show that NF-kappa B activity dynamics in individual cells are not asynchronous and instead resemble the dynamics of the population average in contrast to studies of cells overexpressing p65-EGFP. Molecular & Cellular Proteomics 8:433-442, 2009.