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Background: The present study was conducted to compare various methods of disinfection of impression materials such as glutaraldehyde, ultraviolet (UV) radiation, and autoclave. Materials and Methods: The present study was conducted on eighty alginate impression materials which were disinfectant with 2.2% glutaraldehyde, UV radiation, and autoclave. The pre- and postbacterial count was assessed. Results: The mean premicrobial contamination in Group I was 362.2 106 colony-forming unit (CFU)/ml, in Group II was 306.4 106 CFU/ml, and in Group III was 336.2 106 CFU/ml. The mean postmicrobial contamination in Group I was 65.2 106 CFU/ml, in Group II was 76.7 106 CFU/ml, and in Group III was 28.4 106 CFU/ml. Conclusion: The authors found that autoclave proved to be better in terms of reduction in bacterial colonies as compared to glutaraldehyde and UV radiation.