The proliferation of pharmaceuticals and chemicals and the increasing manufacture of medical goods have made an assessment of their potential toxicological risks to man and the environment indispensable. Hence, a wide range of tests has been developed in order to generate data on the harmful effects of chemicals and pharmaceuticals on organisms. Initially, toxicological data were commonly collected from tests involving animals; however, on account of ethical objections, they are gradually being replaced by in vitro cytotoxicity tests. In this work, a new in vitro screening method for the determination of cytotoxicity, based on a novel electrochemical bioactivity sensor system, was implemented. The evaluation was based on current-time curves of a potentiostatic measurement proportional to the reduction of mediator molecules by mammalian cells. Depending on the number of reduced mediator molecules used by the cells, a current signal is produced, which provides information about the viability of living cells. By adding a toxic test substance to the cell, a reduction in the current signal can be observed, depending on the cytotoxicity of the substance. It is possible to quickly create a specific cytotoxic curve and to determine the corresponding inhibitory concentrations (IC50 value). First tests were performed on three mammalian cell lines and eight model compounds using this electrochemical measurement system. The IC50 values obtained corresponded well with the toxicity determined with an established reference cytotoxicity assay (MTT test) and with data reported in the literature.