Statistical procedures for estimating the detection limit and determination limit of the Ames Salmonella mutagenicity assay

Novel and flexible procedures for estimating the detection limit as well as the determination limit of the Ames mutagenicity assay were proposed to evaluate the genetoxicity of a water sample. The accumulated data under the test conditions of TA 100-S9 by our group were taken as examples and analyzed to estimate the detection limit and the determination limit. The detection limit was estimated at 1.7 as the MR value when duplicate plates were used in the negative control test. However, it decreased to 1.4 as the MR level when quadruple plates were used in the negative control test. Therefore it was found that the sensitivity of the Ames mutagenicity assay was improved very easily by increasing the number of plates for the negative control test from two to four. The application of the conventional twofold rule to the data obtained with the strain TA100 was considered too conservative. The determination limit was regarded at 2.2 as the MR value under the following conditions: (a) quadruple plates were used in the negative control test; (b) three dose-steps including negative control step were designed at regular intervals; and (c) duplicate plates were used for each dose-step. It was proved by comparing data of two students that the detection limit and the determination limit estimated in this study were considered acceptable to any well trained students.