Ulcerative keratitis is a common disease in horses which may cause blindness. To prevent secondary bacterial and fungal infections and promote quick re-growth of the epithelial layer, different treatment approaches have been employed. This study aimed to examine the effects of advanced platelet-rich fibrin (A-PRF) gel on the healing process of experimentally induced corneal ulcers in donkeys. Nine healthy adult donkeys were used for the study. The donkeys were divided into two groups: the control group, where no medication was applied to the corneal ulcer, and the A-PRF gel group, where A-PRF gel was applied once a day on specific days after ulcer induction. The healing process was evaluated through various examinations and analyses. The results demonstrated that the A-PRF gel group showed significant improvement in the corneal ulcer area, with epithelial and stromal regeneration. At day 35, about 60% of the A-PRF group showed negative fluorescein uptake. Additionally, fewer complications were observed during the healing process compared to the control group. In conclusion, A-PRF gel is an important and safe therapeutic option for controlling ocular surface infection and promoting corneal healing. We recommend using A-PRF gel as an alternative approach, avoiding eyelid suturing, and minimizing corneal irritation.© 2023. The Author(s).