in up to independent replicates per concentration. The retention of neutral red by living cells was determined photometrically as described by Seiler and coworkers. 5 The viability of the exposed cells was expressed as the percentage of the negative contro and the data were plotted as concentration “response Tacrolimus curves. Regression analysis was performed using SigmaPlo and extract concentrations inducing 0, 5 and 0 mortalities were calculated accordingly.et assay Theet assay was performed under alkaline conditions according to Singh 6 with modi ations detailed by Keiter and Kosmehl and coworkers. 1 RTL cells were exposed to all genotoxins and extracts for 8 h. MNNG and the extracts were additionally tested using an exposure time of 4 h.
Usual in order to reach high effects in sh cel it is appropriate to use an exposure time of 4 h. 7 In the literatu primary cells from liver and gills of zebra sh were exposed to different genotoxic agents for and 0 h, respective and almost identical effects were found. 8 As a positive control for genotoxici Genistein inhibitor RTL cells were exposed to UVC light for min applying a dose of mW cm . For analys the Olive tail moments as well as the percentage of DNA in the tail of randomly selected nucleoids were measured on two replicate slides. 9 For percent tail D the proportion of DNA that remained in the nucleus is J. Environ. Monit. Downloaded by New York University on 0 March Published on 8 February on pubs.rsc | doi: / E F View Online subtracted from the total nucleolar DNA of a single cell.
The Olive tail moment is calculated by multiplying the distance between the center of the head and the center of the tail by the proportion of total DNA in the tail. This parameter gives values in arbitrary units depending on the software used. Neverthele in addition to Sympatol 94075 percent tail D the tail moment was deemed appropriate for the GeneTEQ concept because all data used for calculation in this study were collected using the same software: Komet ” . Forparison of tail moment and percent tail D GeneTEQs were calculated using both parameters. The induction factor was calculated by dividing the median tail moment at each concentration by the median tail moment of the corresponding negative controls; the IFpensates for variability between different runs. 1 Moreov the concentrationdependent induction factors were calculated according to Seitz buy Hordenine and coworkers.
CDIs were calculated using tail moment as well as percent tail DNA. For statistical interpretati data were tested for normality with the ShapiroWilk test a in the case of lack of normal distributi analyzed with Kruskal “Wallis oneway analysis of variance on ranks. In the case of signi ant differences between grou a posthoc test according to Dunn was run to organizing center identify groups that differed signi antly. GeneTEQ calculation In order to obtain GeneTEQs in relation to the maximum effect of the referencepound MNNG after 4 h and 8 h of exposu the percentage of Olive tail moments and standard deviations of the sample in the maximum effect of the referencepound MNNG was calculated. For percent tail D the same approach was applied. Since a log transformation of effect data enhances their statistical power 2 and since it ismon to take the logarithm of concentration.