OECD 442D: KeratinoSens™ assay

The KeratinoSens™ test represents the second key event in the skin sensitization adverse outcome pathway, the activation of keratinocytes in the epidermis. 

The ARE-Nrf2 luciferase test method uses an immortalized adherent cell line derived from HaCaT human keratinocytes. The cell line is stably transfected with a plasmid containing a luciferase gene under the transcriptional control of the SV40 promoter, fused with the ARE, from a gene known to be up-regulated by contact sensitizers. 

The luciferase signal reflects the activation by sensitizers of endogenous Nrf2 dependent genes and the dependence of the luciferase signal in the recombinant cell line on Nrf2 has been demonstrated. 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 substances. 

The data may be used as part of an integrated approach to testing and assessment (IATA) to support the discrimination between skin sensitizers and non-sensitizers for the purpose of hazard classification and labeling.