A dual-sensing platform is proposed based on multi-walled carbon nanotubes/Prussian blue-functionalized polypyrrole nanowire array (PPY/MWCNTs/PB). Highly aligned PPY nanowire arrays were electrochemically prepared on the surface of glassy carbon electrodes, which were doped with MWCNTs/PB nanocomposites. The nanomaterial combines the characteristics of the PPY nanowires (high conductivity and large specific surface area) and MWCNTs/PB (excellent catalytic performance and intrinsic redox activity). Owing to the nanowire microstructure and outstanding electrical properties, the PPY/MWCNTs/PB nanowire arrays show excellent electrocatalysis of the reduction of hydrogen peroxide and facilitate the construction of a high-performance biosensing platform for microRNA (miRNA). A linear relationship between analytical signal and concentration of hydrogen peroxide and miRNA was obtained in the range 5 to 503 µM (1.4-5.1 mM) and 0.1 pM to 1 nM, and detection limits of 1.7 μM and 33.4 fM, respectively. This new supersensitive sensing platform has broad application prospects of biomolecule and other analyte determination in drug, biomedical, plant protection, and environmental analysis. Prussian blue/multi-walled carbon nanotubes functionalized polypyrrole nanowire arrays (PPY/MWCNTs/PB) were prepared by a facile one-step electrochemical method. PPY/MWCNTs/PB nanowire arrays show excellent electrocatalysis of the reduction of HO and facilitate the construction of a high-performance biosensing platform for microRNA.