© 2020 Elsevier Inc.Objective: The purpose of this video presentation is to demonstrate the effect of intraoperative dilute topical fluorescein in perilympatic fistula diagnosis and localization. Materials and methods: Explorative tympanotomy was performed for the diagnosis, localization and repair of the fistula in the patient who had a pre-diagnosis of perilymphatic fistula. Topical fluorescein was applied intraoperatively to localize the defect. Result: A clear change of color was distinguished from yellow to green leading to diagnosis of the perilymphatic fistula and also showed the origin of the fistula. Conclusion: Topical application of dilute fluorescein is a convenient and effective tool in the diagnosis and localization of perilymphatic fistula.