Validation of the cytosensor for in vitro cytotoxicity studies

The Cytosensor() microphysiometer was used to continuously monitor perturbations in metabolic rates of the human liver cell line ATCC-CCL-13 when exposed to each of 10 drugs. The effects of exposure to one concentration for 24 hr or to sequential increasing concentrations for 4 hr, and recovery after drug removal, were compared. Paracetamol (acetaminophen) and ethanol were used to establish the assay protocols and determine reversibility of drug effect. All drugs produced concentration-and time-dependent reduction in acidification rate following 24 hr exposure, which may be due to decreased number of viable cells and/or lowered metabolic rates of the live cells. The degree of irreversible inhibition of acidification rate was used as an index of cell death and the IC(50) values for the 10 drugs were comparable to those produced in the same cell line by a fluorescence assay using Calcein AM stain (r = 0.991), that fluoresces only in live cells, as well as the [(3)H]thymidine uptake assay (r = 0.976). There was also excellent correlation (r = 0.958) between IC(50) values of 24 hr exposure obtained from the Cytosensor with the 10 drugs and their published human lethal blood concentrations. An advantage of this new methodology over other in vitro assays is that it allows the determination of time points at which reversible change becomes irreversible.