Real-Time Sensing of Single-Ligand Delivery with Nanoaperture-Integrated Microfluidic Devices

W. Elliott Martin, Ning Ge, Bernadeta R. Srijanto, Emily Furnish, C. Patrick Collier, Christine A. Trinkle, Christopher I. Richards

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

The measurement of biological events on the surface of live cells at the single-molecule level is complicated by several factors including high protein densities that are incompatible with single-molecule imaging, cellular autofluorescence, and protein mobility on the cell surface. Here, we fabricated a device composed of an array of nanoscale apertures coupled with a microfluidic delivery system to quantify single-ligand interactions with proteins on the cell surface. We cultured live cells directly on the device and isolated individual epidermal growth factor receptors (EGFRs) in the apertures while delivering fluorescently labeled epidermal growth factor. We observed single ligands binding to EGFRs, allowing us to quantify the ligand turnover in real time. These results demonstrate that this nanoaperture-coupled microfluidic device allows for the spatial isolation of individual membrane proteins while maintaining them in their cellular environment, providing the capability to monitor single-ligand binding events while maintaining receptors in their physiological environment. These methods should be applicable to a wide range of membrane proteins.

Original languageEnglish
Pages (from-to)3858-3867
Number of pages10
JournalACS Omega
Volume2
Issue number7
DOIs
StatePublished - Jul 31 2017

Bibliographical note

Funding Information:
The authors acknowledge A. Sorkin (Professor, Department of Pharmacology, University of Colorado) for EGFR−GFP plasmid through Addgene (plasmid 32751). This work was funded through the Human Frontiers Science Program (RGY0081/2014) and the National Institute of Health DA 038817.

Publisher Copyright:
© 2017 American Chemical Society.

ASJC Scopus subject areas

  • Chemistry (all)
  • Chemical Engineering (all)

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