Abstract The development of a highly selective and sensitive H2O2 sensor, based on the electro-deposition of Prussian blue (PB) onto screen-printed carbon electrodes (SPCEs) modified by benzethonium chloride (BZTC), is described. This methodology provides a time-efficient method for producing stable films in the presence of a BZTC concentration which was optimized with respect to electrochemical and electroanalytical properties. Some parameters such as the amount of PB deposited, film thickness, reversibility, permeability, stability, electrocatalytic properties and sensitivity to H2O2 were considered in order to select the optimal sensor. Under optimal conditions (2 mM BZTC), the surface coverage and apparent diffusion coefficient for K+ displayed values of 6.4 ± 0.2 × 10−8 mol cm−2 and 5.2 × 10−11 cm2 s−1, respectively, one order of magnitude higher than without surfactant. The catalytic rate constant for the optimized film was 2.8 × 103 M−1 s−1 which was in good agreement with data found in the literature. BZTC(2 mM)/PB operating at ∼0 V vs. SCE displayed the highest H2O2 sensitivity (1.07 ± 0.03 A M−1 cm−2, n = 5) reported in the literature to date for PB-modified SPCEs, and showed an excellent limit of detection (