What is the official description of skin corrosion?
Skin corrosion refers to the production of irreversible damage to the skin manifested as visible necrosis through the epidermis and into the dermis, following the application of a test chemical [as defined by the United Nations (UN) Globally Harmonized System of Classification and Labeling of Chemicals (GHS)]. (from OECD TG 431)
Which is the current OECD Guideline for skin corrosion testing?
The current OECD Test Guideline 431 / OCDE Ligne Directrice 431 "OECD GUIDELINES FOR THE TESTING OF CHEMICALS; In vitro skin corrosion: reconstructed human epidermis (RHE) test method" was adopted on 28 July 2015. They are available on the OECD homepage and here:
Here, we provide an extract of this Test Guideline. If you need more detailed information, please consult the full guideline as PDF.
updated Test Guideline 431 provides an
in vitro procedure allowing the
identification of non-corrosive and corrosive substances and mixtures in
accordance with UN GHS. It also allows a partial sub-categorization of
This Test Guideline addresses the human health endpoint skin corrosion. It 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 the human skin , i.e. the epidermis. This Test Guideline was originally adopted in 2004 and updated in 2013, 2014 and 2015 to include a set of Performance Standards (PS) (Annex 1) for the assessment of similar and modified RhE-based test methods, in accordance with the principles of Guidance Document No. 34. Other updates comprise the addition of two test methods using the RhE models SkinEthic RHE and epiCS (previously named EST-1000), and the possibility to use the methods to support the sub-categorisation of corrosive chemicals.
validated test methods using commercially available RhE models are included in
this Test Guideline. Prevalidation studies, followed by a formal validation
study for assessing skin corrosion have been conducted for two of these
commercially available test methods, EpiSkin™ Standard Model (SM) and EpiDerm™
Skin Corrosivity Test (SCT) (EPI-200) (designated the Validated Reference
Methods – VRMs). The outcome of these studies led to the recommendation that
the two VRMs mentioned above could be used for regulatory purposes for
distinguishing corrosive from non-corrosive substances and that the EpiSkin could moreover be used to support sub-categorization of corrosive substances. Two other commercially available in vitro skin corrosion RhE test methods have shown similar results to the EpiDerm VRM according to PS-based validation. These are the SkinEthic RHE and epiCS (previously named EST-1000) that can also be used for regulatory purposes for distinguishing corrosive from non corrosivesubstances. Post validation studies performed by the RhE model producers in the years 2012 to 2014 with a refined protocol correcting interferences of unspecific MTT reduction by the test chemicals improved the performance of both discrimination of C/NC as well as supporting sub-categorisation of corrosives.
This Test Guideline addresses the in vitro skin corrosion component of the tiered testing strategy recommended within TG 404 for dermal corrosion/irritation assessment. It allows the identification of non-corrosive and corrosive substances and mixtures in accordance with the UN GHS. This Test Guideline further allows the sub-categorization of corrosive substances and mixtures into optional Category 1A, in accordance with the UN GHS, as well as a combination of Categories 1B and 1C. A limitation of this Test Guideline is that it does not allow discriminating between skin corrosive sub-categories 1B and 1C in accordance with the UN GHS due to the limited set of well known in vivo corrosive Cat 1C chemicals.
basis of the overall data available, the Test Guideline is applicable to a wide
range of chemical classes and physical states including liquids, semi-solids,
solids and waxes.
While this Test Guideline does not provide adequate information on skin irritation, it should be noted that OECD TG 439 specifically addresses the health effect skin irritation in vitro and is based on the same RhE test system, though using another protocol. For a full evaluation of local skin effects after a single dermal exposure, it is recommended to follow the sequential testing strategy as appended to TG 404. This testing strategy includes the conduct of in vitro tests for skin corrosion (such as described in this Test Guideline) and skin irritation before considering testing in living animals. [...]
How does the test work?
[...] The test
chemical is applied topically to a three-dimensional RhE model, comprised of
non-transformed, human-derived epidermal keratinocytes, which have been
cultured to form a multilayered, highly differentiated model of the human epidermis. [...]
[...] The RhE test method is based on the premise that corrosive chemicals are able to penetrate the stratum corneum by diffusion or erosion, and are cytotoxic to the cells in the underlying layers. Cell viability is measured by enzymatic conversion of the vital dye MTT [...] into a blue formazan salt that is quantitatively measured after extraction from tissues. Corrosive chemicals are identified by their ability to decrease cell viability below defined threshold levels. [...]
How can I translate viability data into substance classification?
exposure time points (t=3 and
|Prediction to be considered|
|< 50% after 3 min exposure||Corrosive:|
Optional Sub-category 1A
50% after 3 min exposure AND|
< 15% after 60 min exposure
A combination of optional Sub-categories 1B and 1C
50% after 3 min exposure AND |
≥ 15% after 60 min exposure
How does this work in practice?
This case study illustrates skin corrosion testing results and the classification of test chemicals for three test substances as an example.
Case study skin corrosion
Substance A is a solid substance, which is applied as a powder (after grinding) topically onto the epidermis.
|Case A||Subset 1||Subset 2|
|Exposure times||3 min||1 h|
|Viability||73 %||39 %|
|Classification||Non-Corrosive; No Category|
Substance B is a creamy substance, which is applied with a brush topically onto the epidermis.
|Case B||Subset 1||Subset 2|
|Exposure times||3 min||1 h|
|Viability||61 %||9 %|
|Classification||Corrosive; Cat 1B/C|
Substance C is a liquid substance, which is applied by pipetting topically onto the epidermis.
|Case C||Subset 1||Subset 2|
|Exposure times||3 min||1 h|
|Viability||18 %||7 %|
|Classification||Corrosive, Cat 1A|
Which methods are validated and accepted?
validated methods are included in the OECD TG 431:
epiCS, EpiDerm, EpiSkin and SkinEthic SM.
The validation study for the epiCS method for skin corrosion testing revealed very good results for discrimination of corrosive from non-corrosive chemicals (Corrosive versus Non-Corrosive). This test method can also be used for sub-categorisation of corrosives.
|epiCS test method|
|Overall Accuracy||92,4 %|
In parallel, the prediction models of the test methods were under investigation in order to improve predictivity. The resulting paper "Two novel prediction models improve predictions of skin corrosive sub-categories by test methods of OECD Test Guideline No.431" by DESPREZ et al. was published in Toxicology in vitro (2015).
What do I need for skin corrosion testing?
We have developed a specific kit that you can use for in vitro skin corrosion testing. In addition you need laboratory material which are standards in cell culture laboratories.
- epiCS (tissues)
- epiCS Culture Medium - to culture the tissues - for three medium changes (sufficient for skin corrosion testing)
- epiCS MTT-Assay Medium - for MTT-Assay
- 6-well plates
- Lot specific epiCS certificate of analysis
We provide competent swift technical and scientific support by telephone or e-mail.
How can I compare test performances in different labs?
designed a validation kit for skin corrosion to conduct a multicenter
study to validate between laboratory reliability.
Two (blinded) test substances are tested at your site(s) and at our ISO9001:2008 certified laboratories. Testing is carried out at the same time on the same lot of tissues to minimize material variations between the sites. Test results are compared and in case of concordant results a certificate is issued by CellSystems to the qualified user. This study can be carried out with one or more external laboratories at the same time
Who should participate?
a) Beginners - to prove that the test
method is carried out correctly following the SOP.
b) Routine users - to prove on a routine basis the reproducible performance of the technicians.
For further information, please download the PDF below
SOP epiCS in vitro Skin Corrosion, v6.0, 2015
- UN (2013), United Nations Globally Harmonized System of Classification and Labeling of Chemicals (GHS), Fifth revised edition, UN New York and Geneva.
- OECD (2004), In vitro skin model. OECD Guideline for the Testing of Chemicals No. 431, OECD, Paris.
- ICCVAM (2004), Recommended Performance Standards for In Vitro Test Methods for Skin Corrosion. NIH Publication Number 04-4510. Research Triangle Park, NC: National Institute of Environmental Health Sciences.
- OECD (2005), Guidance Document on the Validation and International Acceptance of New or Updated Test Methods for Hazard Assessment. OECD Series on Testing and Assessment No. 34, OECD, Paris.
- EC-ECVAM (1998), Statement on the scientific validity of the EpiSkinTM test (an in vitro test for skin corrosivity), issued by the ECVAM Scientific Advisory Committee (ESAC10), 3 April 1998.
- EC-ECVAM (2000), Statement on the application of the EpiDermTM human skin model for skin corrosivity testing, issued by the ECVAM Scientific Advisory Committee (ESAC14), 21 March 2000.
- Hoffmann et al., (2005), Epidermal-skin-test 1000 (EST-1000)-A new reconstructed epidermis for in vitro skin corrosivity testing. Toxicol. in Vitro 19, 925-929.
- Kandárová, H. et al. (2006), Assessment of the human epidermis model SkinEthic RHE for in vitro skin corrosion testing of chemicals according to new OECD TG 431. Toxicol.in Vitro 20, 547–559.
- EC-ECVAM (2009), ESAC statement on the scientific validity of an in-vitro test method for skin corrosivity testing: the EST-1000, issued by the ECVAM Scientific Advisory Committee (ESAC30), 12 June 2009.
- OECD (2013),Summary Document on the Statistical Performance of Methods in OECD Test Guideline 431 for Sub-categorisation. OECD Series on Testing and Assessment No. 190. OECD, Paris.
- OECD (2015), OECD Guidelines for the testing chemicals; In vitro skin corrosion: reconstructed human epidermis (RHE) test method.