An intelligent, autocatalytic, DNAzyme biocircuit for amplified imaging of intracellular microRNAs

DNAzymes hold great promise as transducing agents for the analysis of intracellular biomarkers. However, their low intracellular delivery efficiency and limited signal amplification capability (including an additional supply of cofactors) hinder their application in low-abundance biomarker analysis. Herein, a general strategy to design an intelligent, autocatalytic, DNAzyme biocircuit is developed for amplified microRNA imaging in living cells. The DNAzyme biocircuit is constructed based on a nanodevice composed of catalytic hairpin assembly (CHA) and DNAzyme biocatalytic functional units, sustained by Au nanoparticles (AuNPs) and MnO₂ nanosheets (CD/AM nanodevices). Once the CD/AM nanodevices are endocytosed by cells, the MnO₂ nanosheets are reduced by intracellular glutathione (GSH), which not only releases the different units of the DNAzyme circuit, but also generates the cofactor Mn²⁺ for DNAzyme autocatalysis. The intracellular analytes could trigger the coordinated cross-activation of CHA and autocatalytic DNAzymes on AuNPs, enabling reliable and accurate detection of miRNAs in living cells. This intelligent autocatalytic multilayer DNAzyme biocircuit can effectively avoid signal leakage and obtain high amplification gain, expanding the application of programmable complex DNA nanocircuits in biosensing, nanomaterial assembly, and biomedicine.