Single Cell MicroRNA Analysis Using Microfluidic Flow Cytometry

Wu, Meiye; Piccini, Matthew; Koh, Chung-Yan; Lam, Kit S.; Singh, Anup K.

doi:10.1371/journal.pone.0055044

Single Cell MicroRNA Analysis Using Microfluidic Flow Cytometry

Journal Article · Wed Jan 30 00:00:00 EST 2013 · PLoS ONE

DOI:https://doi.org/10.1371/journal.pone.0055044· OSTI ID:1627582

Wu, Meiye ^[1]; Piccini, Matthew ^[2]; Koh, Chung-Yan ^[2]; Lam, Kit S. ^[3]; Singh, Anup K. ^[4]

Sandia National Lab. (SNL-CA), Livermore, CA (United States). Dept. of Biotechnology and Bioengineering; Univ. of California, Berkeley, CA (United States). Biochemistry and Molecular Biology Graduate Group; DOE/OSTI
Sandia National Lab. (SNL-CA), Livermore, CA (United States). Dept. of Biotechnology and Bioengineering
Univ. of California, Berkeley, CA (United States). Biochemistry and Molecular Biology Graduate Group; Univ. of California, Davis, CA (United States). Dept. of Hematology and Oncology
Univ. of California, Berkeley, CA (United States). Biochemistry and Molecular Biology Graduate Group

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.

View Accepted Manuscript (DOE)

Research Organization:: Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division

Grant/Contract Number:: AC04-94AL85000

OSTI ID:: 1627582

Journal Information:: PLoS ONE, Journal Name: PLoS ONE Journal Issue: 1 Vol. 8; ISSN 1932-6203

Publisher:: Public Library of ScienceCopyright Statement

Country of Publication:: United States

Language:: English

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