Single-Cell Measurements of IgE-Mediated FcεRI Signaling Using an Integrated Microfluidic Platform

Liu, Yanli; Barua, Dipak; Liu, Peng; Wilson, Bridget S.; Oliver, Janet M.; Hlavacek, William S.; Singh, Anup K.

doi:10.1371/journal.pone.0060159

Title: Single-Cell Measurements of IgE-Mediated FcεRI Signaling Using an Integrated Microfluidic Platform

Journal Article · Wed Mar 27 00:00:00 EDT 2013 · PLoS ONE

DOI:https://doi.org/10.1371/journal.pone.0060159· OSTI ID:1321718

Liu, Yanli ^[1]; Barua, Dipak ^[2]; Liu, Peng ^[1]; Wilson, Bridget S. ^[3]; Oliver, Janet M. ^[3]; Hlavacek, William S. ^[2]; Singh, Anup K. ^[1]

Sandia National Lab. (SNL-CA), Livermore, CA (United States). Biotechnology and Bioengineering Dept.
Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Theoretical Biology and Biophysics Group. Theoretical Division. Center for Nonlinear Studies
Univ. of New Mexico, Albuquerque, NM (United States). Dept. of Pathology. Cancer Center

Heterogeneity in responses of cells to a stimulus, such as a pathogen or allergen, can potentially play an important role in deciding the fate of the responding cell population and the overall systemic response. Measuring heterogeneous responses requires tools capable of interrogating individual cells. Cell signaling studies commonly do not have single-cell resolution because of the limitations of techniques used such as Westerns, ELISAs, mass spectrometry, and DNA microarrays. Microfluidics devices are increasingly being used to overcome these limitations. In this paper, we report on a microfluidic platform for cell signaling analysis that combines two orthogonal single-cell measurement technologies: on-chip flow cytometry and optical imaging. The device seamlessly integrates cell culture, stimulation, and preparation with downstream measurements permitting hands-free, automated analysis to minimize experimental variability. The platform was used to interrogate IgE receptor (FcεRI) signaling, which is responsible for triggering allergic reactions, in RBL-2H3 cells. Following on-chip crosslinking of IgE-FcεRI complexes by multivalent antigen, we monitored signaling events including protein phosphorylation, calcium mobilization and the release of inflammatory mediators. The results demonstrate the ability of our platform to produce quantitative measurements on a cell-by-cell basis from just a few hundred cells. Finally, model-based analysis of the Syk phosphorylation data suggests that heterogeneity in Syk phosphorylation can be attributed to protein copy number variations, with the level of Syk phosphorylation being particularly sensitive to the copy number of Lyn.

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Research Organization:: Los Alamos National Laboratory (LANL), Los Alamos, NM (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States)

Sponsoring Organization:: USDOE; National Inst. of Health (NIH) (United States)

Contributing Organization:: Univ. of New Mexico, Albuquerque, NM (United States)

Grant/Contract Number:: AC52-06NA25396; AC04-94AL85000; P50GM085273

OSTI ID:: 1321718

Report Number(s):: LA-UR-12-23884

Journal Information:: PLoS ONE, Vol. 8, Issue 3; ISSN 1932-6203

Publisher:: Public Library of ScienceCopyright Statement

Country of Publication:: United States

Language:: English

Citation Metrics:

Cited by: 21 works

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