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The SkinEthic Skin irritation Test (SIT) can be used to predict the acute skin irritation potential of chemicals/substances.

The method uses the SkinEthic reconstructed human epidermis model and involves the topical application of a chemical for 42 minutes followed by 42 hours incubation and then determination of cell viability via the MTT assay.

The method and protocol have received regulatory approval as a full replacement for the in vivo (rabbit) skin irritation test under an EC test method regulation as method B.46 and OECD test guideline No. 439.

The results of the validation study, the peer review of ESAC and our recommendations can be found on TSAR, the Tracking System for Alternative methods towards Regulatory acceptance.

Read more about the SkinEthic Skin Irritation Test (SIT) on TSAR

Skin irritation

[collapsed]Acute skin irritation is a local reversible inflammatory response or damage to skin that occurs following the application of a chemical/substance. This is in contrast to skin corrosion, which is normally considered to be non-reversible damage to skin.

The potential of a chemical to cause skin irritation is an important consideration with respect to procedures for the safe handling, packaging and transportation of chemicals.[/collapse]

SkinEthic Skin irritation Test (SIT)

[collapsed]The SkinEthic Skin irritation Test (SIT) is an in vitro method that is based on a reconstructed human epidermis model of the same name.

The model consists of normal human keratinocytes that have been cultured for 17 days on an inert polycarbonate filter at an air-liquid interface.

At day 17 the model consists of a highly differentiated and stratified epidermis comprising of a number of layers and a functional stratum corneum (the main skin barrier layer), making it morphologically comparable to skin tissue in vivo.

The test involves exposure of the model to a test chemical/substance for 42 minutes.

After removal of the chemical, the model is then incubated for a further 42 hours. At this point cell viability is established through the MTT assay and compared to the value obtained for a negative control (exposure to phosphate buffered saline).

A chemical is classed as an irritant if ≤50% of cells are viable or a non-irritant if >50% cells are viable.

The current protocol (available on TSAR) is based on the EpiSkin skin irritation test method and is validated according to EURL ECVAM performance standards and OECD requirements.[/collapse]

Animal testing replacement

[collapsed]The test method has received regulatory approval as a replacement for the in vivo Draize rabbit skin irritation test.

An OECD test guideline (No 439) has previously been adopted that includes this method as validated for replacing the in vivo equivalent.[/collapse]

Validation study

[collapsed]The SkinEthic Skin irritation Test (SIT) underwent a catch-up validation study to benchmark the method against the EpiSkin skin irritation test method and corresponding EURL ECVAM performance standards.

A total of 20 test substances (10 irritants and 10 non-irritants) were assessed in the model, three times in three separate laboratories under GLP-like conditions. An independent statistical analysis was then performed to assess intra- and inter-laboratory reproducibility.

As well as the primary outcome based the MTT assay for cell viability, a secondary outcome, IL-1α cytokine release, was also assessed.[/collapse]

Validation study outcomes

[collapsed]Correct predictions of the skin irritation potential of the tested chemicals resulted in a sensitivity of 90% and specificity of 80%. Overall accuracy was 85%.

The additional IL-1α cytokine release outcome did not improve this level of accuracy.[/collapse]

EURL ECVAM recommendations

[collapsed]ESAC concluded that the performance of the SkinEthic Skin irritation Test (SIT) method met the criteria to be considered sufficiently accurate and reliable for the prediction of skin irritation and non-irritation potential of chemicals. Specifically this was in comparison to the validated and accepted EpiSkin reference method.

Subsequently the method received regulatory approval as a replacement of the in vivo Draize rabbit skin irritation test method.

This was in the form of EC test method regulation as method B.46 and OECD test guideline No. 439.[/collapse]