OECD 492B | The Reconstructed Human Cornea-like Epithelium (RhCE) Test Method for Eye Hazard Identification 

The OECD 492B test is an internationally recognized in vitro method for evaluating the potential of chemicals (substances and mixtures) to cause eye irritation or serious eye damage. It is widely used for regulatory safety assessment and identifies on its own substances requiring classification under the UN Globally Harmonized System (GHS) for ocular hazards:

• Category 1 (Serious eye damage classification); H318
• Category 2 (Eye irritation classification); H319
• No Category (No classification required)

The SkinEthic™ Human Corneal Epithelium (HCE) Time-to-Toxicity (TTT) test, as described in OECD 492B, is the latest addition to the NAMs (New Approach Methodologies / Non-animal Alternative Methods) among the OECD guidelines for eye hazard testing. It represents an improvement over OECD Test Guidelines 437 and 492, which together allow the identification of chemicals "not requiring classification" (No Category) or chemicals with serious eye damage potential but could not distinguish between Category 2 and Category 1 substances. And even more, the HCE-TTT test fully replaces animal testing methods, such as the Draize test.

Test Principle

The three-dimensional reconstructed human cornea-like epithelium (RhCE) model from SkinEthic™ closely mimics the histological and biochemical properties of the human corneal epithelium. Cell viability, which correlates with tissue health, is quantified by the enzymatic reduction of the tetrazolium salt (MTT) to a formazan dye. Reduced viability/cytotoxicity indicates a potential ocular hazard. The test employs multiple exposure times (2 for solids, 3 for liquids) to improve the accuracy and predictability of eye irritation categories (Time-To-Toxicity approach).

Test Scope

The test involves the measurement of MTT reduction on the SkinEthic™ tissue model following different exposure times with the test item in comparison to tissues treated with  a negative control and a positive control in order to evaluate the eye hazard potential of a test chemical based on its ability to induce cytotoxicity.

Test Variant

For strongly colored test items absorbing light in the same wavelength range as the formazan dye where photometric evaluation is not possible due to interference, we offer an analytical approach using High-Performance Liquid Chromatography (HPLC) to reliably quantify the formazan production.

Please contact us for more information.