The National Vaccination Programme (NVP) is an exceptionally effective intervention in the field of public health, aiming at eliminating mortality caused by diseases for which there are available vaccines. Background: The study aims at recording the factors that determine parents’ level of knowledge about vaccines and their attitudes in Greece. Methods: A total of 2995 parents were asked to fill in a specially structured questionnaire. The study was conducted in 107 Athens kindergartens. Results: 58.8% of the parents demonstrated a satisfactory level of knowledge and alertness with respect to vaccination and NVP. Parents’ age, educational level, professional status, nationality, religion and sources of information contribute to parental awareness over the importance of vaccination. Conclusions: Knowledge and timely information constitute the most important factors that positively affect the attitudes of parents with regard to vaccines and NVP. In-school education of parents may ensure their consistency in NVP implementation.