Multiple microRNAs analysis based on DNAzyme cascade DNA fiber barcodes for cell typing

Accurate tumor subtyping through multiplex miRNA profiling is critical for precision medicine but remains challenging due to limitations in current methods, including cross-reactivity, cost, and stability. Herein, we present a DNAzyme-powered DNA fiber barcode platform that combines G-quadruplex (G4)/hemin catalysis, miRNA-responsive displacement, and polydopamine (PDA)-mediated fluorescence modulation for sensitive and specific miRNA detection. The system operates via a target-dependent “switch”: in the absence of target miRNAs, G4/hemin DNAzymes catalyze dopamine (DA) oxidation into PDA that efficiently quench fluorescence through Förster resonance energy transfer (FRET); when target miRNAs are present, they competitively binding to the capture strands on DNA fibers with higher affinity, displacing the G4 structures, inhibiting DNAzyme formation and preserving the fluorescence signal, with fluorescence intensity showing a positive correlation with target concentration. This mechanism enables dual qualitative and quantitative analysis with a broad linear range (50 pM-100 nM) and low detection limits (5.50 pM). Key advantages include stable, tunable G4/hemin transduction, enhanced kinetics and signal-to-noise through synergistic displacement-quenching, and multicolor barcoding for one-pot multiplex miRNA detection. Validated against qPCR with high concordance, this platform overcomes existing technical barriers to enable robust miRNA-based cell typing classification and precision diagnostics.