Test of the Month!
KeratinoSens™ with an XCellR8 twist!
Dr Carol Treasure
Our festive choice for Test of the Month is one of our most established methods – KeratinoSens! This test assesses the human skin sensitisation potential of a wide range of substances. It’s a fully approved OECD Test Guideline, and we can provide the test to full GLP for submissions under REACH, CLP and other global regulations. We also provide it as a non-regulatory screen to maximise accessibility to human-relevant skin sensitisation data across a wider cross-section of industry, helping to ensure that skin sensitisation potential of an ingredient is flagged at an early stage in NPD.
Every one of our tests had a unique story behind it. Read on to find out how we’ve added our unique XCellR8 twist to this well-known method, in line with our mission to accelerate the world’s transition to 100% animal-free testing!
First – a bit of context
KeratinoSens was developed by Dr Andreas Natsch and his team at the Swiss fragrance company, Givaudan. The international validation trial was published in 2013. It was part of a wider industry push at the time, to fully elucidate the skin sensitisation Adverse Outcome Pathway (AOP) and to establish non-animal tests for each of the Key Events. Methods were subsequently approved for Key Event 1 (the binding of the chemical to skin peptides – this step is known as the molecular initiating event), Key Event 2 (activation of keratinocytes in the epidermis) and Key Event 3 (activation of the immune response via the skin’s dendritic cells). KeratinoSens covers Key Event 2 and is included in OECD’s recently published Defined Approaches on Skin Sensitisation (Guideline 497) – an important guidance document on how to use the tests and other information sources to complete a safety assessment for the skin sensitisation endpoint.
How does the test work?
As we’ve said, KeratinoSens addresses Key Event 2, which takes place in the cells of the epidermis (keratinocytes). It induces inflammatory responses as well as gene expression associated with specific cell signalling pathways. The KeratinoSens™ cell line is a human keratinocyte cell line, containing a luciferase gene that is linked with a gene known to be up-regulated by contact sensitisers. In this test, a sensitising test item switches on the luciferase-linked gene, meaning that luciferase itself is also produced by the cells. Quantitative measurement (by luminescence detection) of luciferase activity is then carried out using a standard substrate. The prediction model correlates the luciferase signal with sensitisation potential. A test item inducing luciferase activity by more than or equal to 1.5-fold over the untreated control level is classified as Positive (Sensitiser). A test item causing less than 1.5-fold induction is classified as Negative (Non-Sensitiser). We test a range of concentrations and there must be a dose-responsive effect for the prediction to be valid. While the test is designed for a binary (yes/no) classification for hazard identification purposes, testing a concentration range can provide additional useful information about where the threshold is crossed, enhancing the value of the data for risk assessment purposes.
So, what’s XCellR8’s unique twist?
When we set up KeratinoSens at XCellR8 in 2014, we knew we needed to adapt it to completely animal product free (APF) conditions, in line with our APF policy, which runs through everything we do. This has the scientific advantage of maximising the human relevance of our tests, along with the ethical benefits of removing the use of animal-derived components. With amazing support from UK-based global cosmetics brand Lush, we adapted the KeratinoSens cell line to culture in human serum, eradicated the other animal-based reagents used in the original method, and ran the recommended panels of reference chemicals to ensure that our adaptation gave robust results. But the practical steps were only the beginning! Since we were deviating from the original method, we needed to get the adaptation approved by OECD so that we could support cosmetics ingredient suppliers and other companies who wanted to use it in regulatory submissions. We took our work to the OECD Expert Working Group on skin sensitisation, and after an in-depth review process, the test was accepted! It’s now included within OECD Test Guideline 442D (Appendix 1A, Annex 2). We also published the project in the ALTEX journal to share the validation data.
The whole process led to a new debate at OECD level about the importance of animal product free approaches to in vitro Test Guidelines, the challenges and opportunities around the use of human reagents and chemically defined systems. As a small team, we’re proud to have made an impact and so excited to see the growing support for APF approaches across industry and other stakeholders.
Adoption by industry
We’ve been providing KeratinoSens to industry for a decade now. XCellR8 was one of the first labs to provide the test, and we’ve built up a wealth of experience. We’re conducting the method regularly for cosmetics ingredients suppliers as well as product formulators to support new product development (NPD) pipelines. Our reach goes well beyond cosmetics too: we work with chemical companies across diverse sectors from agriculture and ship building, aviation and motor sport, to household products and medical devices. Alongside KeratinoSens, we can advise on a broader testing strategy for skin sensitisation depending on the nature of the chemical or product, in line with OECD Guideline 497.
Test of the month
To celebrate our “test of the month”, we’re offering special discounts on KeratinoSens, for orders placed throughout the month of December. If you’re not ready to test yet, you can place your order now to secure your lab slot and take advantage of the lower price. Discounts vary according to the number of items to test. So, whether you’re trying out this test for the first time, or already familiar with the method, find out more on our website, or contact us to arrange a conversation about how KeratinoSens can support your animal-free safety assessment process!
Email info@x-cellr8.com
Call +44 (0)1925 607 134.
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