Development of a new opacitometer for the bovine corneal opacity and permeability (BCOP) assay

The bovine corneal opacity and permeability (BCOP) assay is an in vitro eye irritation test that relies on the measurement of induced opacity and permeability evaluated in isolated bovine corneas. Routinely, opacity is measured by an OP-KIT opacitometer which provides a centre-weighted reading of light transmission through the corneal tissue. However, this may underestimate opacity that develops as spots or heterogeneous opaque areas on the periphery of isolated cornea. Consequently, these findings question the reliability of the obtained data when opacity is manifested in a non-homogeneous manner. In addition, the BCOP test is only recommended to identify corrosive and severe eye irritants, and has not proven as sensitive in distinguishing among mild and moderate eye irritants. Within this context, a new opacitometer was developed which uses an adjustable laser beam in combination with a calibrated photometer. This work focused on a technical optimisation and describes the optical characteristics of the new device. The laser-based opacitometer allowed the analysis of the complete corneal surface, showed improved sensitivity to detect subtle changes in corneal transparency and was able to detect more efficiently opaque spots located along the sides of the excised corneas.