Water supplied through dental unit waterlines (DUWLs) has been shown to contain high number of bacteria. To reduce the contamination of DUWLs, it is essential to develop effective disinfectants. It is, however, difficulty to obtain proper DUWL samples for studies. The purpose of this study was to establish a simple laboratory model for reproducing DUWL biofilms. The bacteria obtained from DUWLs were cultured in R2A liquid medium for 10 days, and then stored at −70℃. This stock was inoculated into R2A liquid medium and incubated in batch mode. After 5 days of culturing, it was inoculated into the biofilm formation model developed in this study. Our biofilm formation model comprised of a beaker containing R2A liquid medium and five glass rods attached to DUWL polyurethane tubing. Biofilm was allowed to form on the stir plate and the medium was replaced every 2 days. After 4 days of biofilm formation in the laboratory model, biofilm thickness, morphological characteristics and distribution of the composing bacteria were examined by confocal laser microscopy and scanning electron microscopy. The mean of biofilm accumulation was 4.68×10⁴ colony forming unit/cm² and its thickness was 10~14 µm. In our laboratory model, thick bacterial lumps were observed in some parts of the tubing. To test the suitability of this biofilm model system, the effectiveness of disinfectants such as sodium hypochlorite, hydrogen peroxide, and chlorhexidine, was examined by their application to the biofilm formed in our model. Lower concentrations of disinfectants were less effective in reducing the count of bacteria constituting the biofilm. These results showed that our DUWL biofilm laboratory model was appropriate for comparison of disinfectant effects. Our laboratory model is expected to be useful for various other purposes in further studies.