OBJECTIVE We previously reported that selective depletion of β-lymphocytes with rituximab, an anti-CD20 monoclonal antibody, slowed decline of b-cell function in recentonset type 1 diabetesmellitus (T1DM) at 1 year. Subjects were followed further to determine whether there was persistence of effect. RESEARCH DESIGN AND METHODS Eighty-seven subjects (aged 8-40 years) were randomly assigned to, and 81 received, infusions of rituximab or placebo on days 1, 8, 15, and 22. The primary outcomedbaseline-adjusted mean 2-h area under the curve (AUC) serum C-peptide during a mixed-meal tolerance test (MMTT) at 1 yeardshowed higher C-peptide AUC with rituximab versus placebo. Subjects were further followed with additional MMTTs every 6 months. RESULTS The rate of decline of C-peptide was parallel between groups but shifted by 8.2 months in rituximab-treated subjects. Over 30 months, AUC, insulin dose, and HbA1c were similar for rituximab and placebo. However, in evaluating change in C-peptide over the entire follow-up period, the rituximab group means were significantly larger as compared within assessment times with the placebo group means using a global test (P = 0.03). Odds ratio for loss of C-peptide to,0.2 nmol/L following rituximab was 0.565 (P = 0.064). B-lymphocytes recovered to baseline values by 18 months. Serum IgG levels were maintained in the normal range but IgM levels were depressed. CONCLUSIONS Like several other immunotherapeutic approaches tested, in recent-onset T1DM, rituximab delays the fall in C-peptide but does not appear to fundamentally alter the underlying pathophysiology of the disease.