Objective To investigate the clinical effect of mitomycin C （MMC） on the corneal endothelium and corneal posterior surface after laser subepithelial keratomileusis （LASEK）. Methods In a continuity prospective double-blind study， 0.02% MMC was used topically for 20 seconds in 33 eyes of 33 patients who had undergone LASEK after excimer laser ablation in the Fudan EENT Hospital. The other eye of these patients without MMC treatment was used as the control. Three and 6 months after surgery， the changes in the curve and elevation of the corneal posterior surface， and the density and variation coefficient of corneal endothelial cells were observed to evaluate the MMC toxicity． Data were analyzed using a paired t test. Results In the MMC-treated eyes， 3 and 6 months after the operation， the average corneal endothelium cell density was 3353.3±290.3 cells/mm2 and 3407.6±298.4 cells/mm2； the variation coefficient was 13.36%±6.29% and 13.48±4.34%. In the control eyes 3 and 6 months after the operation， the average corneal endothelium cell density was 3258.7±284.6 cells/mm2 and 3339.2±213.4 cells/mm2； the variation coefficient was 14.52%±7.85% and 14.06%±6.50%. The changes in the elevation and curve of the corneal posterior surface in the MMC treated eyes were -0.67±2.26 μm and -0.52±2.67 μm and 0.01±0.20 D and 0.01±0.21 D， 3 and 6 months after the operation. The changes in the control eyes were -0.85±2.31 μm and -0.15±3.00 μm and -0.01±0.22 D and 0.05±0.17 D at the respective time points after the operation. There were no significant differences in the above parameters either between preoperative or postoperative data or between these two groups. Conclusion The study showed that the application of 0.02% mitomycin C in LASEK is safe.