In 2006/2007 a large serum bank was established by means of a cross-sectional population-based study. This serum bank will be used to evaluate the Dutch national immunisation programme (NIP) by serosurveillance and additional immunological and epidemiological research. In this paper we describe the design of this population-based cross-sectional serosurvey and report the participation rates as well as general characteristics of the study population. A similar serum bank was collected in 1995/1996. Dutch inhabitants (aged 0-79 years, men and women) were invited from 40 municipalities throughout the country and also from eight additional municipalities known with low vaccination coverage (LVC). An oversampling of the migrant population was performed. Blood samples were obtained from all participants accompanied with extensive information on demographic and epidemiological data, such as vaccination history, risk factors and travelling. In addition, sociodemographic data are available from individuals who declined to participate (non-response survey). Overall 33% of all invitees were included in this study. The serum bank comprises 6386 sera in the nationwide sample including the extra sample of immigrants (n=646) and 1518 sera from the LVC municipalities. The sera will be analysed for antibodies against all NI P antigens but will also be used for other infectious diseases research. Results of this second serosurveillance study will contribute to the discussion whether it is needed to reconsider the schedule and/or the vaccine components of the current National Immunisation Programme.