The glass ionomer cement Vitrebond showed a clear genotoxic effect in the in vitro Mammalian Cell Gene Mutation Test (HPRT Test) with CHO cells as well as in the bacterial umu-test with Salmonella typhimurium TA1535/pSK1002. Both DMSO and Ham's F12 cell culture medium extracts according to ISO 10993-12 (Biological evaluation of medical devices-Part 12: sample preparation and reference materials, Geneva, Switzerland) exhibit a clear genotoxic effect in the umu-test. The effect is independent of the extraction volume in a range from 0.5 to 4 ml Ham's F12 cell culture medium. Subsequent extractions of Vitrebond showed no significant difference in the genotoxic response although weight loss and content of 2-hydroxyethyl-methacrylate dropped significantly. In vivo conditions of Vitrebond were simulated by extractions with artificial and collected human saliva. These extracts showed a clear genotoxic effect in the umu-test, even if only a few seconds of extraction time were applied. In conclusion, sample preparations for genotoxicity testing according to ISO 10993-12 reflect the in vivo conditions of Vitrebond applications. This seems to be mostly due to the hydrophilic nature of the genotoxic ingredients.