Fluorescent miRNA analysis enhanced by mesopore effects of polydopamine nanoquenchers

The combination of fluorophore-labelled single-strand DNA probes and nanomaterial quenchers has shown great potential in miRNA detection. The development of advanced detection systems by understanding and controlling the fluorescence quenching/recovery via nanoquenchers' microstructures and local morphologies is an attractive area warranting further investigations. Inspired by nanopore sequencing, we present a novel miRNA sensing strategy using fluorophore-labeled DNA as probes and a type of large-pore-sized mesoporous polydopamine nanoparticles (MPDA-L, 70 nm in diameter) as fluorescence quenchers. It is revealed that the quenching efficiency of MPDA-L towards the fluorophore labelled on the probe, reached more than 99% at a relatively low particle concentration. Moreover, the mesopores effectively protected the probe DNA from cleavage by DNase I which was used for signal amplification. Sensitive detection of miRNA with a low detection limit of 32-40 pM, as well as a linear detection range of up to 5 nM, was realized by the mesopore effects via a greatly improved differential affinity of ssDNA and the probe-miRNA heteroduplex toward the surface of nanoquenchers. Interestingly, enhanced DLVO (Derjaguin-Landau-Verwey-Overbeek) repulsion generated inside the pore surface by the negative surface-curvature effect correlates with the improved duplex detachment and fluorescence recovery. The developed strategy can be successfully applied to quantify down-regulated let-7a and up-regulated miRNA-21 in different types of cancer cells by using total RNA samples from cell lysate. These findings are expected to inspire strategies and pave a way for utilizing porous nanomaterials for constructing miRNA detection systems.