Dynamic gene expression profiling using a microfabricated living cell array.
We describe the development of a microfluidic platform for continuous monitoring of gene expression in live cells. This optically transparent microfluidic device integrates high-throughput molecular stimulation with nondestructive monitoring of expression events in individual living cells, hence, a living cell array (LCA). Several concentrations of a soluble molecular stimulus are generated in an upstream microfluidic network and used to stimulate downstream reporter cells, each containing a green fluorescence reporter plasmid for a gene of interest. Cellular fluorescence is continuously monitored and quantified to infer the expression dynamics of the gene being studied. We demonstrate this approach by profiling the activation of the transcription factor NF-kappaB in HeLa S3 cells in response to varying doses of the inflammatory cytokine TNF-alpha. The LCA platform offers a unique opportunity to simultaneously control dynamic inputs and measure dynamic outputs from adherent mammalian cells in a high-throughput fashion. This approach to profiling expression dynamics, in conjunction with complementary techniques such as DNA microarrays, will help provide a more complete picture of the dynamic cellular response to diverse soluble stimuli.
Thompson
Deanna M
DM
Department of Surgery, Massachusetts General Hospital, Harvard Medical School, and Shriners Hospitals for Children, Boston, Massachusetts 02114, USA.
King
Kevin R
KR
Wieder
Kenneth J
KJ
Toner
Mehmet
M
Yarmush
Martin L
ML
Jayaraman
Arul
A
D020869:Gene Expression Profiling / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D046210:Microfluidic Analytical Techniques; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D052002:NF-kappa B p50 Subunit; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D011498:Protein Precursors