Single-cell proteomic chip for profiling intracellular signaling pathways in single tumor cells
Edited by Chad A. Mirkin, Northwestern University, Evanston, IL, and approved November 17, 2011 (received for review July 6, 2011)
Abstract
We describe a microchip designed to quantify the levels of a dozen cytoplasmic and membrane proteins from single cells. We use the platform to assess protein–protein interactions associated with the EGF-receptor-mediated PI3K signaling pathway. Single-cell sensitivity is achieved by isolating a defined number of cells (n = 0–5) in 2 nL volume chambers, each of which is patterned with two copies of a miniature antibody array. The cells are lysed on-chip, and the levels of released proteins are assayed using the antibody arrays. We investigate three isogenic cell lines representing the cancer glioblastoma multiforme, at the basal level, under EGF stimulation, and under erlotinib inhibition plus EGF stimulation. The measured protein abundances are consistent with previous work, and single-cell analysis uniquely reveals single-cell heterogeneity, and different types and strengths of protein–protein interactions. This platform helps provide a comprehensive picture of altered signal transduction networks in tumor cells and provides insight into the effect of targeted therapies on protein signaling networks.
Acknowledgments.
We thank Bruz Marzolf and Pamela Troisch for printing DNA microarrays, and the University of California, Los Angeles nanolab for photomask fabrication. This work was funded by the National Cancer Institute Grant 5U54 CA119347 (to J.R.H., principal investigator), The Ben and Catherine Ivy Foundation, the Goldhirsch Foundation, and the Grand Duchy of Luxembourg.
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Cover image: Zebrafish (pictured) are a model organism for studying human disease, although the extent of genetic variation between commonly used laboratory strains is not well understood. Kim H. Brown et al. found a high level of copy number variants (CNVs)—gains and losses of large sequences of DNA—among fish representing three commonly used laboratory strains. About 70% of all identified CNVs appeared to be strain-specific. The authors suggest that the extensive genetic variation within and among strains may have functional effects that could potentially confound studies intended for translation to human diseases. See the article by Brown et al. on pages 529–534. Image courtesy of William T. Lester (Massachusetts General Hospital, Boston) and Charles Lee.
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Submission history
Published online: December 27, 2011
Published in issue: January 10, 2012
Acknowledgments
We thank Bruz Marzolf and Pamela Troisch for printing DNA microarrays, and the University of California, Los Angeles nanolab for photomask fabrication. This work was funded by the National Cancer Institute Grant 5U54 CA119347 (to J.R.H., principal investigator), The Ben and Catherine Ivy Foundation, the Goldhirsch Foundation, and the Grand Duchy of Luxembourg.
Notes
This article is a PNAS Direct Submission.
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The authors declare no conflict of interest.
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