Target-triggered stochastic DNAzyme motors on spherical nucleic acids for simultaneous fluorescence assay of double...

Simultaneous quantifications of multiple miRNAs in the single-sampling system would be conducive to the accurate diagnosis of diseases in contrast with single miRNA analysis. In this work, a stochastic DNAzyme motor on spherical nucleic acids (SNAs) for simultaneous fluorescence assay of double miRNAs was established. Hairpin 1 (H1)-FAM-7a and H1-TAMRA-133a-functionalized magnetic beads (MBs) as SNAs were mixed. Targets (let-7a and miRNA-133a) reacted with two different S1 and S2, triggering the formation of two types of metal DNAzymes. The DNAzymes can further react with H1 stem-loop DNA on SNAs to release the two fluorescent DNA-FAM and DNA-TAMRA fragments in the presence of Mg²⁺. Meanwhile, the DNAzyme as DNA motors were separated from the previous H1 probe to participate the next cycling operations, resulting in the signal amplification toward the simultaneous and sensitive detection of let-7a and miRNA-133a. SNAs with three dimensional nanostructures provided enough space for the operation of DNAzyme walker, promoting the sensitivity of this proposed analytical system. The two mixed SNAs enable one-step and specific quantification of miRNA let-7a and miRNA-133a with lower detection limits of 90.5 fM and 74.9 fM, respectively. Finally, this proposed strategy was employed to simultaneously detect double miRNAs in practical applicability.