The BCOP assay is used in the identification of chemicals that cause no ocular irritation or serious damage. However, this method has not been found to adequately discriminate between mild from moderate ocular irritation (category 2A/2B), based upon the animal data. In this study, we aimed to establish methods for discerning ocular irritation by chemicals. We used the BCOP assay and the fluorescence staining methods based on biomarkers for cellular viability and death. The potential for ocular irritation by 12 chemicals from different UN GHS categories was assessed by the BCOP assay. Cryosections of bovine corneas were obtained. The necrotic nucleus was TUNEL labeled, cytoplasmic f-actin was stained by phalloidin while the nucleus was stained by DAPI. The depth of injury (DOI) was then measured. According to BCOP assay,in vivo data of Draize eye test and DOI,the results showed that category NC irritants caused ≤ 10% epithelial DOI, irritants of category 2B caused >10% epithelial DOI and showed no stromal damage, while category 2A showed damage to the stroma. Based on these results, the GHS prediction model could distinguish between GHS 2A and 2B. Authenticating the viability of BCOP by DOI measurements can provide a more reliable basis for classifying ocular irritants.

中文翻译：

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细胞活力和死亡生物标志物能够使用牛角膜混浊度和渗透性测定评估眼部刺激

BCOP 测定用于鉴定不会引起眼睛刺激或严重损伤的化学品。然而，根据动物数据，尚未发现该方法能够充分区分轻度和中度眼部刺激（2A/2B 类）。在这项研究中，我们旨在建立辨别化学品对眼部刺激的方法。我们使用 BCOP 测定和基于细胞活力和死亡生物标志物的荧光染色方法。通过 BCOP 测定评估了来自不同联合国 GHS 类别的 12 种化学品对眼部刺激的可能性。获得牛角膜的冷冻切片。坏死的细胞核被 TUNEL 标记，细胞质 f-肌动蛋白被鬼笔环肽染色，而细胞核被 DAPI 染色。然后测量损伤深度（DOI）。根据 BCOP 测定，Draize眼试验和DOI的体内数据显示，NC类刺激物引起≤10%的上皮DOI，2B类刺激物引起>10%的上皮DOI且未显示基质损伤，而2A类刺激物对基质造成损伤。基于这些结果，GHS 预测模型可以区分 GHS 2A 和 2B。通过 DOI 测量验证 BCOP 的可行性可以为分类眼刺激物提供更可靠的基础。