Jump to main content
Jump to site search

Issue 7, 2015
Previous Article Next Article

A bead-based microfluidic approach to integrated single-cell gene expression analysis by quantitative RT-PCR

Author affiliations

Abstract

Gene expression analysis at the single-cell level is critical to understanding variations among cells in heterogeneous populations. Microfluidic reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) is well suited to gene expression assays of single cells. We present a microfluidic approach that integrates all functional steps for RT-qPCR of a single cell, including isolation and lysis of the cell, as well as purification, reverse transcription and quantitative real-time PCR of messenger RNA in the cell lysate. In this approach, all reactions in the multi-step assay of a single lysed cell can be completed on microbeads, thereby simplifying the design, fabrication and operation of the microfluidic device, as well as facilitating the minimization of sample loss or contamination. In the microfluidic device, a single cell is isolated and lysed; mRNA in the cell lysate is then analyzed by RT-qPCR using primers immobilized on microbeads in a single microchamber whose temperature is controlled in closed loop via an integrated heater and temperature sensor. The utility of the approach was demonstrated by the analysis of the effects of the drug (methyl methanesulfonate, MMS) on the induction of the cyclin-dependent kinase inhibitor 1a (CDKN1A) in single human cancer cells (MCF-7), demonstrating the potential of our approach for efficient, integrated single-cell RT-qPCR for gene expression analysis.

Graphical abstract: A bead-based microfluidic approach to integrated single-cell gene expression analysis by quantitative RT-PCR

Back to tab navigation

Supplementary files

Publication details

The article was received on 29 Oct 2014, accepted on 10 Nov 2014 and first published on 10 Nov 2014


Article type: Paper
DOI: 10.1039/C4RA13356K
Author version available: Download Author version (PDF)
Citation: RSC Adv., 2015,5, 4886-4893
  •   Request permissions

    A bead-based microfluidic approach to integrated single-cell gene expression analysis by quantitative RT-PCR

    H. Sun, T. Olsen, J. Zhu, J. Tao, B. Ponnaiya, S. A. Amundson, D. J. Brenner and Q. Lin, RSC Adv., 2015, 5, 4886
    DOI: 10.1039/C4RA13356K

Search articles by author

Spotlight

Advertisements