Objective To investigate the vaccination rate and evaluate the immunoprophylaxis effect of hepatitis B vaccine (HepB) in Xishuangbanna Prefecture, in order to set up an evidence-based strategy for prevention and control of hepatitis B. Methods Stratified cluster sampling was used to select healthy people aged 0-34 years from 3 counties (city) in Xishuangbanna Prefecture. Their HepB vaccination history was collected and venous blood was sampled. ELISA was used for qualitative analysis of serum markers hepatitis B surface antigen (HBsAg) and hepatitis B surface antibody (HBsAb). Chi-square test was used to compare positivity rates. Results From 2016 to 2020, there were a total of 1 647 cases of hepatitis B in Xishuangbanna, with an average annual incidence of 35.48/100 000. The rate of timely HepB primary immunization was 47.98% and that of complete immunization was 55.26%. In the 3 956 subjects surveyed, the positivity rates of HBsAg and HBsAb were 2.10% [95% confidence interval (CI): 1.65%-2.54%] and 65.72% (95% CI: 65.24%-67.20%), respectively. The positivity rate of HBsAb in male population was statistically significantly higher than that in female population, and higher in city population than in rural population (χ2=18.35 and 4.48, respectively, both P<0.05). The positivity rates of HBsAg were statistically significantly different among 3 counties (city) (χ2=11.51, P<0.01). Conclusions The HepB immunity in Xishuangbanna Prefecture is weaker among rural population. The weak regions should strengthen HepB immunization to improve the vaccination coverage, strengthen boost immunization, and improve comprehensive prevention and control measures such as screening, diagnosis and antiviral treatment for chronic HBV infected persons.