Electrochemical single-cell gene-expression assay combining dielectrophoretic manipulation with secreted alkaline phosphatase reporter system

Tatsuya Murata, Tomoyuki Yasukawa, Hitoshi Shiku, Tomokazu Matsue

Research output: Contribution to journalArticlepeer-review

73 Citations (Scopus)

Abstract

Scanning electrochemical microscopy (SECM) was used for the analysis of single-cell gene-expression signals on the basis of a reporter system. We microfabricated a single-cell array on an Indium tin oxide (ITO) electrode comprising 4 × 4 SU-8 microwells with a diameter of 30 μm and a depth of 25 μm. HeLa cells transfected with plasmid vectors encoding the secreted alkaline phosphatase (SEAP) were seeded in the microwell at a concentration of 1 cell per well by positive-dielectrophoresis (pDEP). A pDEP pulse of 3.0 Vpp and 1 MHz was applied between the microwell array/ITO electrode and an ITO counter electrode located on the top of the flow-cell assembly of the microdevice. The electrochemical responses of the individual HeLa cells transfected with SEAP were significantly larger than those of the wild-type HeLa cells. The electrochemical response of the transfected single cells was statistically distinguishable from that of wild-type HeLa cells. The size of the wells and the material of the single-cell array were optimized in order to evaluate the tumor necrosis factor α (TNF-α)-induced activation process of nuclear factor kappa B (NFκB) that was used as the model for on-chip monitoring of cellular signal transduction.

Original languageEnglish
Pages (from-to)913-919
Number of pages7
JournalBiosensors and Bioelectronics
Volume25
Issue number4
DOIs
Publication statusPublished - 2009 Dec 15

Keywords

  • Dielectrophoresis
  • Reporter
  • Scanning electrochemical microscopy
  • Single-cell analysis

ASJC Scopus subject areas

  • Biotechnology
  • Biophysics
  • Biomedical Engineering
  • Electrochemistry

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