In this video, we demonstrate the EpiDerm Skin Irritation test (EpiDerm SIT) developed and validated for in vitro skin irritation testing of chemicals, including cosmetic and pharmaceutical ingredients.
I. Tissue conditioning – Day 0
II. Chemical exposure – Day 1
III. Medium exchange – Day 2
IV. MTT Viability Assay – Day 3
V. Assay Quality Controls
Representative Results
Figure 1. Results obtained for 6 test articles, NC and PC control – Part of the automated spreadsheet for calculation of results. Test chemicals which reduced tissue viability below 50% referenced to NC are classified as irritants. The test is optimized to provide results that are sufficiently far from the classification cut-off (50% viability) thus increasing the robustness of the assay.
In this video, we have demonstrated the EpiDerm Skin Irritation test (EpiDerm SIT) developed and validated for in vitro skin irritation testing of chemicals, including cosmetic and pharmaceutical ingredients. When performing this method, it is important to work in aseptic conditions and to strictly follow the validated protocol, since deviation from the protocol may cause different outcomes. Some modifications of the assay are possible, however, changes to the protocol should be discussed directly with the authors.
The only limitation of this method is a possible interference of the test substance with the MTT endpoint. A colored test substance or one that directly reduces MTT (and thereby mimics dehydrogenase activity of the cellular mitochondria) may interfere with the MTT endpoint. However, these test substance are a problem only if at the time of the MTT test (i.e. 42 hours after test substance exposure) sufficient amounts of the test substance are still present on (or in) the tissues. In case of this unlikely event, the (true) metabolic MTT reduction and the contribution by a colored test material or (false) direct MTT reduction by the test material can be quantified by a procedure described in detail in the assay Standard Operation Procedure provided by MatTek Corp.
The authors have nothing to disclose.
The authors would like to thank to ZEBET at the BfR (Germany), BASF SE (Germany), IIVS Inc. (USA), Zet-LSL (Austria) for participation in Multi-center International Validation Study of the Modified EpiDerm SIT (5).