Here we present design and assay methodology of a novel electrochemical biosensor with the view to assess cytotoxic effects of key chemicals. The concept is based on mammalian cells as the biological recognition agent (A549 human lung epithelial cells) and measures changes in cellular enzyme activity (acid phosphatase – AP) following 24 hours exposure. AP catalyses the dephosphorylation of 2‐naphthyl phosphate to 2‐naphthol (determined using chronocoulometry) and is indicative of metabolically active cells. Immobilised living cells exposed to pentachlorophenol, cadmium chloride and nickel chloride exhibited a decrease in AP activity which enabled IC50 (50 % reduction in enzyme activity) values of toxic chemicals to be reliably and conveniently determined using electronic detection. The IC50 values obtained for CdCl2 and NiCl2 (65 and 330 µM) were in agreement with those found using the standard MTT cytotoxicity assay (100 and 350 µM). In the case of pentachlorophenol, the value obtained (60 µM) was lower (MTT assay IC50 value>160 µM) suggesting enhanced sensitivity of the electrochemical assay towards pentachlorophenol. It is envisaged that this device could be exploited in the screening of industrial and environmental toxins and has potential in drug testing applications.