In-situ thermal cycling amplification on the surface of upconversion nanoparticles and enabling multiplexed microRNA assay

Surface modification is the prerequisite for yielding reliable bionanomaterials for a variety of applications including molecular diagnosis. Among a library of nanomaterials, upconversion nanoparticles (UCNPs) have emerged as promising candidates for biomolecule sensing toward disease diagnosis. As is well acknowledged, surface modification of UCNPs is required because the as-synthesized UCNPs are generally covered with organic ligands. However, current surface functionalized UCNPs cannot support in-situ signal amplification to boost the sensing performance. In this work, ultra-stable surface modification has been achieved to render UCNPs endurable to high temperatures, allowing for target microRNA (miRNA)-mediated in-situ thermal cycling click ligation on the UCNPs’ surface. By integrating with facile magnetic separation, the content of the target miRNA can be faithfully reflected by the upconversion fluorescence intensity in the reaction system, enabling a simple way for nucleic acid quantification. This design exhibits a lowest detectable concentration that is one order of magnitude lower than that of the one-step conjugation without amplification, and is proven to work well in biological medium. We have also demonstrated its potential in multiplexed miRNA sensing by using UCNPs of different emitting colors. This work achieves thermal cycling-mediated nucleic acid signal amplification on the surface of UCNPs, providing a new avenue for providing UCNP-based biosensors.