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Alternatives to Laboratory Animals - ATLA

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Experience with the HET-CAM Method in the Routine Testing of a Broad Variety of Chemicals and Formulations

Arnhild Schrage, Armin O. Gamer, Ben van Ravenzwaay and Robert Landsiedel

Data on eye irritation are generally needed for the hazard identification of chemicals. For the routine testing of a broad variety of chemicals and formulations, we have used the Hen’s Egg Test-Chorioallantoic Membrane (HET-CAM) method. In the course of a tiered-testing strategy, and due to the lack of global regulatory acceptance of the HET-CAM method, we have also performed the Rabbit Eye Irritation test according to OECD Test Guideline 405. Of the 145 substances tested, 76% were classified as non-irritant/mild irritant and 13% were identified as irritant in vivo, according to the EU classification system (61% and 28%, respectively, with the GHS classification). The remaining 11% were severe irritants in vivo, based on the irreversibility of the effects and not due to sufficiently high irritation scores in the three days following application. The retrospective analysis revealed that the overall accuracy of the HET-CAM assay was 65% and the overall rates of false-negatives (FN) and false-positives (FP) were 50% and 33%, respectively. The HET-CAM assay was sufficiently specific (few FP) for water-soluble substances, but failed to identify nearly all the severe irritants within this group. In contrast, it was highly sensitive (no FN) for non-soluble and oil-soluble substances, but the specificity for this group was rather low. Therefore, we conclude that the HET-CAM assay is not useful in our tiered-testing strategy for eye irritation testing. However, for water-insoluble substances, it might be applicable in combination with another in vitro method, provided that regulatory acceptance is gained.

Full text pdf 38(1), 39–52