The fluorescent indicator pyranine is suitable for measuring stromal and cameral pH in vivo

Three pH-sensitive fluorescent indicators (fluorescein, bis-carboxyethyl-carboxyfluorescein and pyranine) were studied to determine their usefulness as probes for the living cornea and anterior chamber. Of the three dyes, pyranine had properties which render it almost ideally suited for this purpose. Adequate concentrations are reached in the cornea and anterior chamber after topical administration; measurements can be made by fluorophotometry for many hours. The pH is calculated by measuring the ratio of fluorescent intensity at two excitation wavelengths, I463/I404, a measurement which is dependent on pH but independent of the concentration of the fluorophore and other variables which can alter the intensity. In the rabbit eye, pyranine in the cornea and anterior chamber was observed to undergo easily measurable changes in fluorescent ratios associated with lid closure and contact lens wear, indicating its sensitivity to mild changes in pH.