Definition of a new intralaboratory limit value for a positive UDS assay in vitro

In our laboratory, we routinely perform the unscheduled DNA synthesis (UDS) assay in vitro using rat primary liver cells and autoradiographical evaluation. The fixed limit of +5 nuclear net grains (NNG) for a positive UDS response no longer seems to be appropriate because of the advanced techniques that are now used. Considering our historical data (mean vehicle control NNG of 45 experiments = -1.5; highest observed mean NNG value in 126 control slides = +1.63 NNG) we will use a mean value of +2 NNG as our new intralaboratory limit value for a positive UDS assay, independent of the statistical evaluation. Experiments with a mean NNG count in the vehicle control of > +0.5 will be rejected as invalid and repeated since mean values of > +0.23 NNG were never reached. Additionally, the NNG data obtained will be subjected to a newly proposed statistical method which includes both the actual and the mean historical control NNG value. First, the sum of the actual and the mean historical control value is divided by two. Secondly, the calculated control value is pairwise compared with the treated groups by the one-sided Dunnett's test. This might be a helpful tool to evaluate a weak or equivocal UDS response.