Figure 4. Detection of exogenous NO released from DETA-NO/AM in human corneoscleral segments using a fluorescent NO-indicator. A: Increase in DAF-FM fluorescence intensity in quadrants of human corneoscleral segments after treatment with DETA-NONOate/AM (exogenous nitric oxide [NO]-donor) compared with vehicle-treated controls. Quadrants of human donor corneoscleral segments from each eye (n = 4 per group) were pretreated with intracellular NO-indicator DAF-FM dye (10 μM) and then treated with DETA-NONOate/AM (20 μM) or vehicle (0.1% dimethyl sulfoxide [DMSO]) at 37 °C for 30 min. Images were taken using fluorescence microscopy at 100X magnification (Scale bar = 50 μm). B: Quantification of DAF-FM fluorescence intensity per unit area (IntDen/μm²) in DETA-NO/AM and vehicle-treated corneoscleral segments using ImageJ analysis. Data are expressed as means ± standard error of the mean (SEM); n = 4 for each group; *** p<0.001; Two-tailed unpaired Student t test.