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Mol Cell Proteomics. 2009 Mar;8(3):433-42. doi: 10.1074/mcp.M800291-MCP200. Epub 2008 Oct 24.

High content cell screening in a microfluidic device.

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1
Department of Biomedical Engineering, The Johns Hopkins University, Baltimore, Maryland 21218, USA.

Abstract

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-kappaB 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.

PMID:
18953019
PMCID:
PMC2649807
DOI:
10.1074/mcp.M800291-MCP200
[Indexed for MEDLINE]
Free PMC Article
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