However, the obtained results from the in vitro assays unexpectedly shown these substances NOT as corrosive as was expected. To address this apparent data inconsistency and confirm

our suspicion that the RhE models are possibly not suitable for these groups of fatty amine derivatives, some substances were selected for a confirmatory in vivo skin corrosion study in rabbits. The comparison of the results from both the in vitro and the in vivo studies are presented here. In vitro skin corrosion assay makes use of reconstructed human epidermis (RhE) obtained from human derived non-transformed epidermal keratinocytes which closely mimics the histological, morphological, biochemical and physiological properties of the upper parts of Gefitinib the human skin, i.e. the epidermis. Testing of the different substances is done in different labs, applying either EpiDerm™ (EPI-200), or EpiSkin™. Both assays are based on the same principles, but differ in various details.

However, both assays have been validated and approved by ECVAM, and their implementations in the respective labs have been proven to provide reliable and consistent results. OECD 431 provides in its annex 3 a comparison between these assays. Corrosive activity is measured by comparing cell viability after exposure with that of the control in an MTT assay. Possible MTT assay interference by the test substance needs to be assessed. For some substances a limited non-specific reduction was observed which was subtracted from the ODs Cyclopamine research buy of the

test substance treated viable tissues. Duplicate tissues were treated with the test material for different exposure. Additional duplicate tissues were treated with DOK2 the positive and negative control materials. All cultures are subsequently incubated for 3 min, 1 h, and (in case of EpiSkin™ assays) also for 4 h. At the end of the treatment period, the tissues are washed and assessed for tissue viability (MTT assay). Results are expressed as percentage of viability compared to negative control. The test substance is considered to be corrosive to skin: – if the viability after 3 min exposure is less than 50%, or The test substance is considered to be non-corrosive to skin: – if the viability after 3 min exposure is greater than or equal to 50% and the viability after 1 h exposure is greater than or equal to 15%. The test substance is considered to be corrosive to skin: – if the viability after 3 min exposure is less than 35% (Cat. 1A), or The test substance is considered to be non-corrosive to skin: – if the viability after 4 h exposure is ⩾35%. Since severe effects could not be excluded, a stepwise exposure regime was used in which the first animal was treated in a stepwise fashion with three patches, thereby minimizing unnecessary animal harm to acceptable levels. This animal received of 0.