Single-cell proteomic chip for profiling intracellular signaling pathways in single tumor cells
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.
Additional Information© 2012 by the National Academy of Sciences. Edited by Chad A. Mirkin, Northwestern University, Evanston, IL, and approved November 17, 2011 (received for review July 6, 2011). Published online before print December 27, 2011. 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. Author contributions: Q.S., L.Q., and J.R.H. designed research; Q.S., L.Q., F.G., R.F., Y.S.S., and D.G. performed research; Q.S., W.W., L.H., P.S.M., and J.R.H. analyzed data; and Q.S., W.W., L.H., P.S.M., and J.R.H. wrote the paper. The authors declare no conflict of interest.
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Supplemental Material - Appendix.pdf