Objective: Bacterial adhesion is an important parameter in the etiology of primary caries formation in Preventive Dentistry. The aim of the present study was to investigate the effect of surface coating by all-in-one adhesives on inhibition of biofilm attachment on root dentin surface. Methods: Bovine root dentin was sliced and ground with 600-grit SiC. The dentin surfaces were single-coated with one of three all-in-one adhesives; Hybrid Bond (Sun Medical), G-Bond (GC) and Clearfil Tri-S Bond (Kuraray Medical) according to the manufacturer's instructions. The dentin surface without coating was remained as a control. The specimens were stored in saliva for 24 hrs, and then artificial biofilm was grown on the specimens inside the artificial mouth system using freshly cultured Streptococcus mutans, Actinomyces naeslundii, and Lactobacillus casei suspended in phosphate buffer saline (PBS), heart infusion with 1% sucrose and PBS for 24 hrs at 37° C. Each sample with biofilm was transferred into microtubules containing 500 µl of PBS and vibrated with a vortex for 20 s to differentiate detached biofilm (suspended in PBS) and retained biofilm (remained on the samples). Turbidity and Glucan measurements were finally performed to evaluate the amount of retained and detached bacteria on the samples after in vitro biofilm attachment. Data were analyzed statistically using one-way ANOVA & Turkey's HSD (p=0.05). Results: All the coated specimens showed significantly lower susceptibility to biofilm attachment compared with non coated dentin (p<0.05). There was no significant difference in the susceptibility to biofilm attachment among the coated groups. Conclusion: Surface coating by the all-in-one adhesives could inhibit biofilm attachment on root dentin surface. Supported by the grant for COE Program FRMDRTB at TMDU.