Introduction: Azathioprine is effective for maintenance of remission in Crohn's disease, however, duration of efficacy and the dose response relationship has not been fully evaluated. Aims: To investigate whether patients kept in remission by azathioprine treatment for >2 years benefit from further treatment, and to explore dose-response relationship. Patients and methods: In an open 12-month trial, patients with inactive Crohn's disease after >2 years (median 37 months) of azathioprine treatment were randomized to azathioprine withdrawal or continued treatment. Primary end point was relapse defined as: (i) Crohn's disease activity index rise ≥ 75, and Crohn's disease activity index >150 or (ii) disease activity requiring intervention. Results: Of 29 patients, 28 completed the observation period or relapsed. Eleven of 13 patients (85%) continuing azathioprine remained in remission compared with seven of 15 (47%) observed without azathioprine (P = 0.043). In patients who had been treated with azathioprine >1.60 mg/kg/day the difference was even more pronounced, eight of nine (89%) vs. four of 12 (33%) respectively (P = 0.017). Conclusions: Patients with Crohn's disease in remission after >2 years of continuous azathioprine treatment will benefit from further continued treatment. Further controlled studies with azathioprine doses <2.0 mg/kg/ day are needed.