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• Protective levels of aHBs were lower than the expected ones (45.9%). • Protective levels of aHBs in vaccinated children at birth were low (56.1%). • Protective levels of aHBs in vaccinated children at 11 years were higher (75.2%). • Vaccination campaigns against HBV should focus on people born in 1992 or before. Universal vaccination is the most effective strategy to control hepatitis B virus (HBV) infection. In Argentina, vaccination against HBV was incorporated in year 2000 for newborns and in 2003 for 11 years old children. However, there is a paucity of data about protection levels against HBV infection. The aim of this work was to determine the prevalence of seroprotective anti-HBs antibodies (aHBs) in Argentina. Serum samples negative for HBsAg and anti-HBc from 132 children born after year 2000 and 762 blood donors, older than 18 years, from five centers across the country, were analyzed for aHBs. Titers ≥10 mIU/mL were observed in 74/132 children (56.1%) and 336/762 (44.1%) in blood donors. The median age for blood donors was 33.9 (23–43); from them, 210 (27.6%) were born after 1992 and, therefore, were catch-up by vaccine implementation at 11 years old age. Donors born in 1992 or before showed a significantly lower frequency of protection (32.2%) compared to donors born after 1992 (75.2%), p < 0.0001. In addition, significant differences were observed in the status of seroprotection between different participating centers (p = 0.024). Implementation of HBV vaccine in 2000 and 2003 implied an overall increase of the aHBs seroprotective rates, with a particularly adequate response in children vaccinated at 11 years old age. The observed results suggest that population born in 1992 or before is currently the most susceptible. Consequently, it would be advisable to become aware of the risk of transmission in this age group and to stress this population vaccination campaigns. [ABSTRACT FROM AUTHOR]