Quantitative evaluation to predict the eye irritancy of chemicals: Modification of chorioallantoic membrane test by using trypan blue

Abstract

The chorioallantoic membrane (CAM) test using fertile hen's eggs has been reported as a promising alternative method to predict the eye irritancy of chemicals. However, the scoring system of the original CAM test is not satisfactory because of its lack of objectivity and quantitativeness. Furthermore, the test requires skill when the effects of test substances are to be graded. To overcome these disadvantages, a more objective means of judging the injurious effects of chemicals was examined as follows: after treatment with the test chemical, the CAM was stained with trypan blue and the amount of pigment adsorbed was measured spectrophotometrically. The amounts of trypan blue adsorbed with the CAM showed a good correlation with the in vivo eye irritation test scores ( r = 0.89). The findings suggest that the trypan blue staining method is very simple and reproducible, and thus would be a promising alternative method for predicting the eye irritancy of chemicals. However, further studies will be required to validate this test method with a wide variety of chemicals and formulations.