Nanoamplicon Comparator for Live-Cell MicroRNA Imaging.

As an investigative tool, live-cell imaging requires superior probe design to guarantee imaging quality and data validity. The ability to simultaneously address the robustness, sensitivity, and consistency issues in a single-assay system is highly desired, but it remains a largely unsolved challenge. We describe herein a probe-design strategy called a nanoamplicon comparator (NAC) and demonstrate its proof-of-concept utility in intracellular microRNA (miRNA) imaging. This novel designer architecture builds upon spherical nucleic acids (SNAs) for robustness, catalytic hairpin assembly (CHA) for sensitivity, and upconversion nanoparticles (UNPs) for consistency. A catalytic circuit comprising a UNP-hairpin-DNA (UNP-HDNA) conjugate and a hairpin-DNA-organic-fluorophore (HDNA-F) conjugate as probe responds to target miRNA and generates the UNP-HDNA-HDNA-F complex as an NAC for quantitative UNP-to-organic-fluorophore-luminescence-resonance-energy-transfer (LRET) imaging against a native UNP-emission reference channel. An imaging application with miR21 shows the ability to monitor miRNA-expression levels across different cell lines and under an external stimulus.