AbstractMercury film microelectrodes were used to determine iron in osteoblast‐like cell culture medium by performing adsorptive cathodic stripping voltammetry in the presence of catechol. The optimized analytical conditions are a catechol concentration of 5.00 × 10−4 mol L−1, pH 8.0 and a collection potential of −1.80 V which eliminates the copper interference. The reoxidation period was limited to the time between switching to −0.10 V and scanning up to the metal peak (ca. −0.40 V). The limit of detection obtained for a 30 s collection time was 1.93 × 10−8 mol L−1 iron. The results achieved by this procedure and using the square‐wave voltammetry technique are accurate and precise. Biological assays were performed to evaluate the mineralization process.