Despite being a major cyanide species in the process water, it is unclear how iron cyanide influences pyritic gold ore flotation as well as how lead ions influence pyritic gold ore flotation in the presence of iron cyanide. This study aims at revealing the interaction of Fe(CN) and lead ions in pyrite flotation to investigate the strong depressing effect of Fe(CN) on pyritic gold ore flotation and the significant activating effect of lead ions on pyritic gold ore flotation in the presence of Fe(CN) using flotation, zeta potential measurement and surface analysis methods. The flotation results showed that upon 5 × 10 mol/L Fe(CN) addition, pyrite recovery drastically decreased from about 51.3% to 8.6%, while the subsequent addition of 9.5 × 10 mol/L lead ions significantly activated pyrite with the recovery increasing from 8.6% to 91%, which demonstrated that Fe(CN) strongly depressed pyrite flotation, while lead ions completely activated pyrite in the presence of Fe(CN). Zeta potential measurement, surface analysis using Cryogenic X-ray photoelectron spectroscopy (Cryo-XPS) and electrochemical impedance spectroscopy (EIS) revealed that Fe(CN) depression was attributed to the chemical adsorption of Fe(CN) on iron sites of pyrite as Prussian Blue (Fe[Fe(CN)]); however, this hydrophilic layer could be covered totally by lead ions which adsorbed on as lead hydroxide/oxide through electrostatic interactions, which resulted in the significant activation effect of lead ions. The results from this study will lead to improved flotation of gold associated with pyrite in gold flotation plants.