BACKGROUND: Diphtheria cases reported in Central Vietnam since 2013 were mainly in children aged 6-15 years, which may reflect an immunity gap. There is little information on population immunity against diphtheria in countries without a school-entry booster dose. We aimed to measure the age-stratified seroprevalence of anti-diphtheria toxoid antibodies, quantify the change in antibody levels in individuals over time, and estimate the length of protective immunity after vaccination in well-vaccinated communities in Vietnam. METHODS: An age-stratified seroprevalence survey among individuals aged 0-55 years was conducted at Nha Trang, Vietnam. The same participants were followed up after two years to quantify the change in antibody levels. IgG was measured using ELISA. The length of protective immunity after vaccination was estimated using a mixed-effect linear regression model with random intercept. RESULTS: Overall seroprevalence was 26% (95%CI:20-32%). Age-stratified seroprevalence was 68% (95%CI:4-11%), 7% (95%CI:4-11%), 12% (95%CI:7-19%), 33% (95%CI:27-40%), and 28% (95%CI:17-43%) among those aged ≤5, 6-15,16-25, 26-35, and 36-55 years, respectively. The antibody levels declined by 47% (95%CI:31-59%) over two years, and the predicted duration of vaccine-derived protective immunity after receiving four doses was 4.3 years (95%CI:3.5-5.3) among participants aged six years or younger. CONCLUSION: Given the low seroprevalence and short period of vaccine protection, a school-entry booster dose (5-7 years) is recommended in Vietnam.