OBJECTIVE: To investigate the long-term stability of regenerative treatment for tympanic membrane perforation (TMP) using gelatin sponge, basic fibroblast growth factor (bFGF) and fibrin glue in a population with TMP closure. MATERIAL AND METHODS: Treatment could be performed up to four times until perforation closure was achieved. Cases in which perforation closure was finally achieved underwent 1 year of follow-up observation. Perforation closure rate was assessed more than 1 year after initial perforation closure, cases of reperforation were examined, and complications were considered. RESULTS: Postoperative observation after 1 year showed complete closure of the TM remained in 45 of the 54 cases (83.3 %). A high rate of perforation closure was thus maintained. Apart from five cases with small perforations, no complications such as dizziness, ear discharge, or cholesteatoma formation were observed. CONCLUSION: These findings suggest the long-term stability of regeneration therapy using gelatin sponge, bFGF, and fibrin glue. This regeneration therapy appears feasible and safe, with a closure rate of TMP after this regeneration therapy equivalent to that of traditional tympanoplasty and myringoplasty.