DNA centipede nanowires drive rapid picomolar microRNA detection in cells and serum.

Accurate and rapid detection of microRNA (miRNA) is essential for clinical diagnostics and molecular biology. Here, we report a DNA centipede nanoprobe (DCNP)-based localized catalytic hairpin assembly (LCHA) strategy for ultrasensitive detection of miR-21 in cells and serum. In this system, DNA nanowires spatially confine two reactive hairpins (HP1 and HP2) to enhance local concentration and accelerate hybridization kinetics, leading to a fivefold sensitivity improvement over free-CHA. The DCNP enables rapid detection within 40 min, with a limit of detection down to 100 pM, and offers robust performance in complex biological fluids without the need for transfection reagents. High specificity is achieved against homologous and mutant miRNAs. Intracellular imaging of miR-21 in MCF-7 cells displays strong fluorescence signals, consistent with qRT-PCR. Clinical serum samples from colon cancer patients and healthy donors are clearly distinguished, yielding a receiver operating characteristic curve with an AUC of 1.00. This LCHA system demonstrates high sensitivity, rapid response, and reliable intracellular delivery, making it a promising candidate for early cancer diagnostics and point-of-care applications.