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Anal Biochem. 2011 May 15;412(2):165-72. doi: 10.1016/j.ab.2011.01.030. Epub 2011 Feb 1.

An enzyme-linked assay for the rapid quantification of microRNAs based on the viral suppressor of RNA silencing protein p19.

Author information

1
Steacie Institute for Molecular Sciences, National Research Council of Canada, Ottawa, Ontario, Canada K1A 0R6.

Abstract

MicroRNAs (miRNAs) are endogenous posttranscriptional regulators found in all metazoa and play crucial roles in virtually all cellular processes. Their aberrant expression has been linked to several diseased states; therefore, techniques capable of sensitive and specific profiling of the miRNA milieu will have significant application in prognostics, diagnostics, and therapeutics. Here we present a method for rapid quantification of miRNA levels using p19, a tombusvirus-encoded suppressor of RNA interference with sequence-independent and size-selective affinity toward 19-bp RNA duplexes. We present a surface plasmon resonance (SPR)-based miRNA sensing method where RNA probes are immobilized on gold surfaces demonstrating p19's utility in recognition of miRNA-bound probes. This allows detection of miRNAs in the low nanomolar range. To increase the sensitivity, a bead-based enzyme immunoassay was performed, and this technique displays a lower detection limit of 1fmol and a linear dynamic range from 1pmol to 1fmol.

PMID:
21284927
DOI:
10.1016/j.ab.2011.01.030
[Indexed for MEDLINE]

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