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PLoS One. 2013;8(1):e55044. doi: 10.1371/journal.pone.0055044. Epub 2013 Jan 30.

Single cell microRNA analysis using microfluidic flow cytometry.

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1
Department of Biotechnology and Bioengineering, Sandia National Laboratory, Livermore, CA, USA.

Abstract

MicroRNAs (miRNAs) are non-coding small RNAs that have cell type and cell context-dependent expression and function. To study miRNAs at single-cell resolution, we have developed a novel microfluidic approach, where flow fluorescent in situ hybridization (flow-FISH) using locked-nucleic acid probes is combined with rolling circle amplification to detect the presence and localization of miRNA. Furthermore, our flow cytometry approach allows analysis of gene-products potentially targeted by miRNA together with the miRNA in the same cells. We demonstrate simultaneous measurement of miR155 and CD69 in 12-O-tetradecanoylphorbol 13-acetate (PMA) and Ionomycin stimulated Jurkat cells. The flow-FISH method can be completed in ∼10 h, utilizes only 170 nL of reagent per experimental condition, and is the first to directly detect miRNA in single cells using flow cytometry.

PMID:
23383050
PMCID:
PMC3559333
DOI:
10.1371/journal.pone.0055044
[Indexed for MEDLINE]
Free PMC Article
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