Background and Objective: In recent years the incidence opportunistic fungual infections has in creased dramatically. One of the most common fungal pathogen Candida dubliniensis, colonization and biofilm formation on the surfaces of medical devices. The aim of this study was to evaluate the inhibitory activity of zinc oxide nanoparticles against C. dubliniensis biofilm formation by the MTT colorimetric method is measured.Materials and Methods: In this study, ZNO nanoparicles using the sol-gel was prepared, size and type of particles, respectively, by scanning electron microscopy (SEM) and X-Ray-Diffraction were determined. The Minimum inhibitory concentration (MIC) of growth dilution method was done by microdilution test.C. dubliensis biofilms (DSY 1024) after 48h incubation at 37oC inhibited the information of nanoparticles ZNO, SDS and Fluconazole drug as a positive control on Candida biofilms was assessed by MTT tetrasolium salt reduction. The collected data using statistical t-test and SPSS software were analyzed.Results: The results of this study showed that a minimum concentration of inhibitor nanoparticles, ZNO, SDS and fluconazole, respectively: 9.25, 0.02, 8 mg/ml. Inhibitory strength of biofilm adhesion in the presence of nanoparticles ZNO, SDS and fluconazole, equivalent to more than twice the concentration of the MIC was determined.Conclusion: In this study, ZNO nanoparticles synthesized by a chemical method indicat that has anti-fungal properties. Therefore, a new way of working for the prevention of biofilm formation of Candida biofilms particularly associated with medical devices to be.