OECD TG 442E In Vitro Skin Sensitization - Human Cell Line Activation Test (h-CLAT)

The standard “OECD TG 442E In vitro skin sensitization – Human cell line activation test (h-CLAT)”, published under the OECD guidelines on responsible business conduct for multinational enterprises (OECD guidelines), is proposed to address the activation of dendritic cells, a key event in the adverse outcome pathway for skin sensitization.

This test guideline addresses human health hazard endpoint skin sensitization following exposure to a test chemical. Skin sensitization refers to an allergic response following skin contact with the test chemical, as defined by the United Nations Globally Harmonized System of Classification and Labeling of Chemicals.

This standard provides four in vitro test methods that address the same key event in the negative result pathway:

• Human cell line activation test
• U937 Cell line activation test
• Interleukin-8 reporter gene test
• Genomic allergen rapid detection for the evaluation of skin sensitizers

All of these methods are used to support the distinction between skin sensitizers and non-sensitizers in accordance with the United Nations Globally Harmonized System of Classification and Labeling of Chemicals. The test methods described in this test guide quantify changes in the expression of a set of genes associated with the activation process of monocytes and dendritic cells (genomic biomarker signature) following exposure to cell surface markers or sensitizers.

The OECD TG 442E standard is a robust in vitro test for identifying potential chemical skin sensitizers with broad applicability and over 90 percent predictive accuracy. This test provides a binary prediction that classifies test samples as skin sensitizers or non-sensitizers. The purpose of this guideline is to determine whether a chemical substance can cause skin sensitization, i.e., allergic contact dermatitis, by activating specific immune responses without performing animal testing.

The human cell line activation test (h-CLAT) is an in vitro (non-animal) test used to evaluate whether a chemical can cause skin sensitization. Skin sensitization is an allergic reaction caused by an overreaction of the immune system to a substance after skin contact. The h-CLAT is designed to model a key biological step in the development of skin sensitization: the activation of dendritic cells (a type of immune cell) by a sensitizing chemical.

This testing method is important because it replaces older animal-based tests. Furthermore, the OECD TG 442E standard is valued because it provides standardized, scientifically validated, and internationally accepted methods for assessing whether chemicals cause skin sensitization, a type of allergic reaction caused by repeated skin exposure.

The h-CLAT method quantifies changes in the expression of cell surface markers associated with the activation process of monocytes and dendritic cells in the human monocyte leukemia cell line THP-1 following exposure to sensitizers. The measured expression levels of dendritic cell cell surface markers are then used to support the distinction between skin sensitizers and desensitizers.

This method is an in vitro assay that measures changes in cell surface marker expression in THP-1 cells of the human monocyte leukemia cell line after 24 hours of exposure to a test chemical. These surface molecules are typical markers of monocyte THP-1 activation and can mimic the activation of dendritic cells, which play a critical role in T cell priming. Changes in surface marker expression are measured by flow cytometry after cell staining with fluorochrome-labeled antibodies. Cytotoxicity measurement is performed simultaneously to assess whether upregulation of surface marker expression occurs at subcytotoxic concentrations. The relative fluorescence intensity of surface markers compared to the solvent/vehicle control is calculated and used in the prediction model to support the distinction between sensitizers and non-sensitizers.

In short, OECD TG 442 is essential to support ethical, reliable and globally consistent safety testing of chemicals and consumer products, particularly in light of evolving regulations and public demand for cruelty-free science.

Our organization, which has been supporting businesses across all sectors for years through a wide range of testing, measurement, analysis, and evaluation activities, has a strong team of employees who closely follow global developments in science and technology and are constantly improving themselves. In this context, we also provide testing services to businesses in accordance with the OECD TG 442E In vitro skin sensitivity - Human cell line activation test (h-CLAT) standard.