First-order toxicity assays for eye irritation using cell lines: Parameters that affect in vitro evaluation

First-order toxicity assays can be used to rapidly screen test agents. Investigators in many laboratories have used cultured cell lines to obtain correlations between first-order assay end-points and in vivo eye irritation (Draize test) for a wide variety of compounds. Since validation is a key step in assay acceptance, it is important to understand which factors alter the responses of cell-line-based assays. In this study we examine: (1) the presence and configuration of a type I collagen gel; (2) the responses of epithelial (Sf-l-Ep) and fibroblast (Sire and 3T3) cell lines; (3) the total glutathione content, ATP content, methionine incor-poration, and neutral red absorption endpoint assays; (4) alco-hol (Cj-Cg), surfactant (Tween 20), and heavy metal (NiCl) test agents; and (5) test agent exposure time (1 to 24 hr). The pres-ence of a collagen gel and the cell type did not significantly affect endpoint assay R50 (test agent concentration that decreases as