Background: Tissue conditioning materials were introduced about 50 years ago. These are soft, resilient, temporary relining materials which by reducing and evenly distributing stresses on the mucosa of the basal seat, have a rehabilitating effect on unhealthy tissue and allow the condition to return to normal health. Denture stomatitis affects 65% of healthy adult mouths. Aim of the study: To compare Fluconazole and Nystatin in tissue conditioner for treatment of denture stomatitis. Materials and methods: The present study was conducted in the Department of Prosthodontics of the Dental institutions. Clinical isolates of Candida albicans were obtained from the Department of Molecular Microbiology of the institute, to use as test organisms for the current study. Antifungal agents, Nystatin and Fluconazole were mixed into tissue conditioner powder at concentrations of 5% wt/wt in a sterile plate. All disks were contaminated with 100 μl of 1 × 106 CFU/ml C. Albicans cell suspension and the cell culture plate were incubated at 35º C on a rotary shaker for 48 hours. The plates were incubated at 37º C for 48 hours and the colonies were enumerated. Results: We observed that that Nystatin 5% solution was the most efficient for inhibiting attachment and colonization of C. albicans (0.22). Fluconazole 5% solution is partially effective efficient for inhibiting attachment and colonization of C. albicans. Conclusion: The incorporation of Nystatin and Fluconazole into tissue conditioners is effective treatment for chronic atrophic candidiasis in denture users. Nystatin is more efficacious as compared to Fluconazole. Key words: Candidiasis, Fluconazole, Nystatin.