In vitro Skin irritation of chemicals (under OECD TG 439 guidelines)
Predicting the cutaneous irritant potential of chemical products represents an important component of the overall safety evaluation programme for new and existing chemicals.
The recently ECVAM validated in vitro protocol offers an excellent In Vitro-to-In Vivo correlation and is a perfect answer to the evolution of the reglementation such as REACH and European Cosmetic directive. According to ECVAM validation, EPISKIN is the only one model accepted for full replacement of animal testing (1)
Carried out in vitro, the SkinEthic reconstructed Epidermis method makes it possible to evaluate the irritant potential of chemical products, without using animal testing.
- The use of a suitable standardized model
The validated method uses a model equivalent to the human epidermis (3). The reproducibility of the model is checked by histological and function tests (4). Compared to in vivo study, the SkinEthic reconstructed epidermis EpiSkin allows use of measurement systems that produces quantitative, reproducible results.
The ECVAM validation of the method has shown its reproducibility within and between laboratories, and demonstrated that it is rapid and economical.
The MTT cytotoxicity assay (5) is a simple robust test for evaluating cellular viability following application of an irritant/toxic agent. The validated method enables evaluation of the viability of epidermal samples after a set time in contact with the product. IL1 alpha measurement in culture media increases the sensibility of the test. Products can thus be classified according to EU and OECD norms (Irritant R38 / Non-Irritant) (6,7).
- 1-Verification of non-interference between the product to be tested and the MTT reagent
- 2-Verification of the compliance of the conditions for transporting tissues
- 3-Application of the product to be tested and (+) and (-) controls
- 4- Elimination of the test product
- 5- Post-incubation of epidermal samples
- 6- Collect of media for IL1-alpha dosage
- 7- Perform the MTT assay
- 8- Verification of the compliance of the (+) and (-) controls
- 9- Application of the predictive classification model with one endpoint (MTT)
|If viability ≤ 50%|| ||Irritant (R38)|
|If viability > 50%|| ||Potentially non irritant|
|If IL1a release ≤ 60pg/ml|| ||Non irritant|
|If IL1a release > 60pg/ml|| || Irritant (R38)|
- 10- Application of the predictive classification model with two endpoints (MTT + IL1-alpha)
| || MTT Test|| MTT Test + IL1a|
Following the results of the studies performed under ECVAM guidelines and controls, the Episkin tissue model has been validated as the only stand alone alternative method to the animal use for skin irritation of chemicals.
Statement on the validity of in vitro tests for skin irritation. ESAC, ECVAM. April, 27th, 2007.
The In Vitro Acute Skin Irritation of Chemicals: Optimisation of the EPISKIN Prediction Model within the Framework of the ECVAM Validation Process. Cotovio, Grandidier, Roguet… ATLA 33, 329-349, 2005.
The ECVAM International Validation study on In Vitro Test for skin Corrosivity. 2. results and Evaluation by the Management Team Fentem, Botham, Curren, Esdaile, Liebsch… Toxicol. In Vitro 12, 483-527, 1998.