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Amplification-free detection of microRNAs via a rapid microarray-based sandwich assay

Amplification-free detection of microRNAs via a rapid microarray-based sandwich assay

Analytical and Bioanalytical Chemistry 409(14): 3497-3505

The detection and profiling of microRNAs are of great interest in disease diagnosis and prognosis. In this paper, we present a method for the rapid amplification-free detection of microRNAs from total RNA samples. In a two-step sandwich assay approach, fluorescently labeled reporter probes were first hybridized with their corresponding target microRNAs. The reaction mix was then added to a microarray to enable their specific capture and detection. Reporter probes were Tm equalized, enabling specificity by adjusting the length of the capture probe while maintaining the stabilizing effect brought about by coaxial base stacking. The optimized assay can specifically detect microRNAs in spiked samples at concentrations as low as 1 pM and from as little as 100 ng of total RNA in 2 h. The detection signal was linear between 1 and 100 pM (R2 = 0.99). Our assay data correlated well with results generated by qPCR when we profiled a select number of breast cancer related microRNAs in a total RNA sample.

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Accession: 059349931

Download citation: RISBibTeXText

PMID: 28349168

DOI: 10.1007/s00216-017-0298-6

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