The aims of this study were to determine anti-HBs positivity in children who had received three doses of hepatitis B vaccine during infancy and to evaluate the factors that may affect the serological status. Local ethics committee approval was obtained at the beginning of the study. The study was carried out between December 2005 and October 2007 among children attending the outpatient clinics of medical school hospital. The study encompassed 912 children (393 female, 519 male; aged 1-5years old) who had been immunized with three doses of intramuscular recombinant hepatitis B vaccine during infancy. All of the children were born to HBsAg negative mothers and did not have any known immune system problems. Sociodemographic characteristics and passive smoking status were gathered by a questionnaire. Anthropometric measurements were taken, and a detailed physical examination was carried out for each child. Blood samples were obtained to check serum HBsAg, anti-HBs and anti-HBc levels by commercial micro-ELISA (Sanofi Diagnostics Pasteur, Sydney) method. Levels of anti-HBs >= 10 mIU/ml were defined as seropositivity. In seronegative children, anti-HBs levels were re-checked 4 weeks after receiving one booster dose of hepatitis B vaccine. Of the children 877 (96.2%) were found anti-HBs positive, while all of them were negative for anti-HBc or HBsAg. Of children 34.8% were 12-23 months; 28.7% were 24-36 months; and 36.5% were 37-60 months-old, and anti-HBs negativity rate was higher in the older age group with a statistically significant difference (1.4%, 3.9% and 4.2%, respectively; p=0.003). Anti-HBs antibodies were found negative in 2.8% of children who were born by vaginal route and in 5.8% of children who were born by cesarean section, the difference being statistically significant (p=0.016). There were no significant differences between anti-HBs seropositivity and gender, working/educational status of the mothers and the presence of smoking parents in the family (p > 0.05). Logistic regression analysis indicated that the factors that affect antibody levels in vaccinated children were the duration of breastfeeding only (4.77 +/- 1.53 months in anti-HBs positives and 3.69 +/- 2.13 months in negatives; p=0.008), birth weight (3328.18 +/- 318 g in anti-HBs positives and 3135.27 +/- 488 g in negatives; p=0.037) and pregnancy parity (anti-HBs was negative in 3.4% of children born from mothers who had < 2 parities, and 8.2% of children born from mothers who had >= 3 parities; p=0.037). The remaining 35 (3.8%) children with undetectable antibody levels became seropositive after one dose of hepatitis B vaccination, with the antibody levels of 100 mIU/ml. This response underlined the presence of immune memory in vaccinated children. The results of this study indicated that almost all 1-5 years old children who had received three doses of hepatitis B vaccine during infancy were protected from hepatitis B virus infection. It was concluded that similar studies should be carried out in different settings.