Objective: Many innovations have recently been implemented in the field of endodontics, often based on a variety of in-vitro/ex-vivo test setups. It was the goal of this research to compare different biofilm models for evaluating the effectiveness of rinsing solutions. Method and Materials: Three different models have been applied in this study, including petri dishes with nutrient medium, arrays of human dentin disks, and split bovine root segments. The susceptibility of biofilms formed by Enterococcus faecalis to commonly used endodontic irrigants was tested. Results: While citric acid 3% did not show an inhibitory effect on solid medium, mean maximum inhibition areolae of 9.8 ± 1.6 mm were found for sodium hypochlorite 3%. Fluorescein solution was shown to penetrate dentin tubules to a depth of 0.5 to 1 mm, indicating that the dentin tubules are not freely accessible, but clotted by E faecalis biofilms. Rinsing root canals with a combination of citric acid, hydrogen peroxide, chlorhexidine, and sodium hypochlorite removed bacterial biofilms from the dentin tubules only to a depth of 0.5 mm, while bacteria in deeper regions were not affected. Conclusion: Standard irrigating solutions inhibit bacterial growth; however, due to the morphology of the tooth, the effect of irrigating media is restricted to the root canal and the adjacent volume of dentin tubules. Results from standardizable diffusion tests seem not to be predictive for clinical performance of irrigating solutions. Keywords: debridement, dentin tubules, endodontic treatment, irrigation