Chitosan (CS) nanofibers were successfully used to modify indium tin oxide (ITO) electrode by electrospinning technique. Then, Prussian blue (PB) nanoparticles were electrodeposited on the CS nanofibers by potentiostatic technique in an acidic solution containing single ferricyanide. By this method, direct synthesis of PB nanoparticles on the nanofibers that were used for modifying electrode came true. Transmission electronic microscopy (TEM) showed that the average size of PB nanoparticles was about 50nm. Selected-area electron diffraction (SAED) showed diffusive diffraction spots, indicating the mosaic structure of the PB nanoparticles on the CS nanofibers. X-ray powder diffraction (XRD) displayed the long-range disorder of CS nanofibers and demonstrated the formation of PB nanoparticles. Results of the scanning electron microscope (SEM) images indicated that the PB nanoparticles could be electrodeposited on the CS nanofibers. The amount of the PB nanoparticles on the CS nanofibers increased with increasing potential value of the electrodeposition. In addition, the cyclic voltammetric response displayed two characteristic redox couples of PB. The modified electrode exhibited electrocatalytic activity towards reduction of H2O2.