Evaluation of the HET-CAM-TSA method as an alternative to the draize eye irritation test

Abstract The Hen's Egg Test-Chorioallantoic Membrane (HET-CAM) method was modified in our laboratory by means of microscopic evaluation, a clear description of the three in vitro endpoints (haemorrhage, lysis and coagulation) and the use of a test substance applicator (TSA). A previous study on 46 chemicals demonstrated the usage of the HET-CAM-TSA assay as a screening test for eye irritancy. In order to extend our database and to come to a more reliable conclusion concerning the use of the HET-CAM-TSA method, a second set of 60 test substances was tested. The in vitro irritation scores (IS) were compared with the in vivo modified maximum average scores (MMAS) calculated 24 hr after instillation. The MMAS irritancy threshold was set at 15.0. The results were analysed according to the Cooper's parameters (specificity, sensitivity and concordance with the Draize test) and the Pearson's correlation coefficient. It was concluded that the HET-CAM-TSA test was a valuable screening test. To compensate for the misclassifications generated, it was also concluded that the HET-CAM-TSA method should be considered as a part of a test battery, together with the Bovine Corneal Opacity-Permeability (BCOP) assay.