To measure and simulate oxygen kinetics during corneal cross-linking at different irradiances with and without supplementary oxygen. Experimental, laboratory study. In de-epithelialized porcine eyes, a femtosecond-laser-generated tunnel was used to place a fiber probe in corneal depths of 100, 200, and 300μm to measure the local oxygen concentration. After riboflavin imbibition, the corneas were irradiated at 3, 9, 18, and 30 mW/cm2 while the oxygen concentration was measured. All experiments were performed under normoxic (21%) and hyperoxic (>95%) conditions. The obtained data were used to identify parameters of a numerical model for oxygen consumption and diffusion. The equilibrium stromal oxygen concentration under atmospheric oxygen at 3 mW/cm2 was 2.3% in 100μm decreasing to <1% in 300μm. With 9, 18, and 30 mW/cm2, no oxygen was available in 200μm, respectively, 100μm or deeper. Using a hyperoxic environment, the concentration was 50% using 3 mW/cm2 in 100μm, decreasing to 40% in 300μm. At 9 mW/cm2, the concentrations were 5%, 3%, and 1% in 100, 200 and 300μm, respectively. Using 18 and 30 mW/cm2, all oxygen was depleted at 100μm; however, oxygen half-lives were longer at 18 mW/cm2 than at 30 mW/cm2. The oxygen model was able to reproduce the experiments and indicated an exponential decay with increasing distance to the anterior surface. Supplementary oxygen increases the oxygen availability during corneal cross-linking. At higher irradiances, supplementary oxygen is beneficial and eliminates the bottleneck of oxygen allowing a potentially more efficient cross-linking. The calibrated numerical model can quantify the spatial oxygen concentration related to different scenarios such as irradiance or environmental oxygen concentration.