Test No. 442D: In Vitro Skin Sensitisation
ARE-Nrf2 Luciferase Test Method
The present Test Guideline addresses the human health hazard endpoint skin sensitisation,
following exposure to a test chemical. Skin sensitisation refers to an allergic response
following skin contact with the tested chemical, as defined by the United Nations
Globally Harmonized System of Classification and Labelling of Chemicals (UN GHS).
This Test Guideline (TG) provides an in vitro procedure (the ARE-Nrf2 luciferase test
method) used for supporting the discrimination between skin sensitisers and non-sensitisers
in accordance with the UN GHS. The second key event on the adverse outcome pathway
leading to skin sensitisation takes place in the keratinocytes and includes inflammatory
responses as well as gene expression associated with specific cell signalling pathways
such as the antioxidant/electrophile response element (ARE)-dependent pathways. The
test method described in this Test Guideline (ARE-Nrf2 luciferase test method) is
proposed to address this second key event. The cell line contains the luciferase gene
under the transcriptional control of a constitutive promoter fused with an ARE element
from a gene that is known to be up-regulated by contact sensitisers. The luciferase
signal reflects the activation by sensitisers of endogenous Nrf2 dependent genes.
This allows quantitative measurement (by luminescence detection) of luciferase gene
induction, using well established light producing luciferase substrates, as an indicator
of the activity of the Nrf2 transcription factor in cells following exposure to electrophilic
test substances. There are currently two in vitro ARE-Nrf2 luciferase test method
covered by this Test Guideline: the KeratinoSensTM test method and the LuSens test
method. Performance standards have been developed to enable the validation of similar
test methods.
Available from June 30, 2022Also available in: French
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