This follow-up study assessed the 2-year clinical and magnetic resonance imaging (MRI) outcomes of patients with multiple sclerosis (MS) originally enrolled in an MRI study conducted at eight centres in south Italy (the South Italy Mobile MRI Project). Of the,597 MS patients recruited at baseline, 391 returned for the follow-up study. Of these, 363 provided 2-year clinical and MRI follow-up data, and 215 were still undergoing treatment with one of four interferon beta regimens: Avonex, 30 mcg intramuscularly once weekly; Betaferon, 250 mcg subcutaneously (sc) every other day; Rebif 22 mcg sc three times weekly (tiw; Rebif 22); or Rebif 44 mcg sc tiw (Rebif 44). Over the 2-year follow-up period, patients receiving the higher dose of Rebif were more likely to remain free from relapses [odds ratio (OR) = 2.23] and from developing both new T2 (OR = 0.15) and new T1 black hole lesions (OR = 0.22), when compared with patients in the Avonex group. Despite some limitations in the trial design, the results from this follow-up study provide helpful clinical and MRI data on the efficacy of interferon beta regimens in MS patients treated in the clinical setting.