Single cell kinase signaling assay using pinched flow coupled droplet microfluidics

Ramesh Ramji, Ming Wang, Ali Asgar S. Bhagat, Daniel Tan Shao Weng, Nitish V. Thakor, Chwee Teck Lim, Chia Hung Chen

Research output: Contribution to journalArticle

Abstract

Droplet-based microfluidics has shown potential in high throughput single cell assays by encapsulating individual cells in water-in-oil emulsions. Ordering cells in a micro-channel is necessary to encapsulate individual cells into droplets further enhancing the assay efficiency. This is typically limited due to the difficulty of preparing high-density cell solutions and maintaining them without cell aggregation in long channels (>5 cm). In this study, we developed a short pinched flow channel (5 mm) to separate cell aggregates and to form a uniform cell distribution in a droplet-generating platform that encapsulated single cells with >55% encapsulation efficiency beating Poisson encapsulation statistics. Using this platform and commercially available Sox substrates (8-hydroxy-5-(N,N-dimethylsulfonamido)-2-methylquinoline), we have demonstrated a high throughput dynamic single cell signaling assay to measure the activity of receptor tyrosine kinases (RTKs) in lung cancer cells triggered by cell surface ligand binding. The phosphorylation of the substrates resulted in fluorescent emission, showing a sigmoidal increase over a 12 h period. The result exhibited a heterogeneous signaling rate in individual cells and showed various levels of drug resistance when treated with the tyrosine kinase inhibitor, gefitinib.

Original languageEnglish (US)
Article number034104
JournalBiomicrofluidics
Volume8
Issue number3
DOIs
StatePublished - 2014
Externally publishedYes

ASJC Scopus subject areas

  • Biomedical Engineering
  • Materials Science(all)
  • Condensed Matter Physics
  • Fluid Flow and Transfer Processes
  • Colloid and Surface Chemistry

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    Ramji, R., Wang, M., Bhagat, A. A. S., Weng, D. T. S., Thakor, N. V., Lim, C. T., & Chen, C. H. (2014). Single cell kinase signaling assay using pinched flow coupled droplet microfluidics. Biomicrofluidics, 8(3), [034104]. https://doi.org/10.1063/1.4878635