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

作者：Nasheri Neda; Cheng Jenny; Singaravelu Ragunath; Wu Peng; McDermott Mark T; Pezacki John Paul^*

来源：Analytical Biochemistry, 2011, 412(2): 165-172.

DOI：10.1016/j.ab.2011.01.030

• MicroRNAs
• Surface plasmon resonance
• Quantification
• Enzyme immunoassay
• p19

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 1 fmol and a linear dynamic range from 1 pmol to 1 fmol.

• 出版日期2011-5-15