Tick-borne encephalitis (TBE) is a serious viral infection, which can lead to permanent neurological sequelae in children. The incidence of TBE disease is increasing in many European countries and is particularly pronounced in some regional populations. Vaccination is the most effective method for preventing TBE disease and is recommended for all those living and working in TBE-endemic areas. Encepur Children is licensed for TBE vaccination in children 1-11 years of age. Following primary vaccination, booster vaccinations are recommended; however, the optimal timing for booster vaccination of children is not known. The aim of this study was to assess the persistence of TBE antibodies in children at 3 and 5 years after their first booster vaccination with Encepur Children and to re-evaluate booster vaccination recommendations. Children 1-11 years of age (n=335) who received primary TBE vaccination according to the rapid schedule (Days 0, 7, and 21) in a previous study received a booster vaccination 12-18 months later, and were invited for follow-up at 3 and 5 years post-booster. TBE antibodies were measured using a virus neutralization test (NT; in-house, Novartis Vaccines) and also using anti-TBE IgG ELISA (Enzygnost, Siemens, Germany). In this analysis, 275 of 278 (99%) subjects and all 190 (100%) subjects who completed the follow-up at 3 and 5 years, respectively, had NT titres > or = 10. Likewise, all 275 of 278 (99%) and 188 of 190 (99%) subjects tested positive by ELISA at 3 and 5 years after the booster vaccination, respectively. Based on serological data, the interval for subsequent booster vaccinations with Encepur Children can be extended from 3 to 5 years after receiving primary vaccination and a first booster vaccination 12-18 months later.