Paper/plastic sterilization pouches are commonly used packaging material for steam sterilization. Reuse of these pouches is a general practice in Thailand despite a single-use recommendation. This study aimed to determine microbial contamination after reusing paper/plastic sterilization pouches in a dental clinic and storage in a closed environment for 6 months. Three hundred and twenty pouches underwent 3 times of clinical use in terms of packaging, autoclave sterilization, handling, and unpacking. A mouth mirror was packed in each pouch to be used in a clinic. After each use, a pouch would be carefully inspected for reusability and undergone packaging, sterilization, handling again. In all steps, sterilization monitoring was rigorously applied. After 3 times of use, a piece of filter paper was placed inside each pouch (instead of a mouth mirror), the pouch was autoclaved and stored in a closed environment for 6 months. Then the filter paper was retrieved for microbial cultivation. A negative control group comprised new pouches containing filter paper without storage and a positive control group comprised pouches with impaired integrity. All samples in both the reuse and the negative control groups had no microbial contamination. All samples in the positive control group showed contamination. These results suggested that reusing paper/plastic sterilization pouches could be a safe practice provided careful monitoring and inspection were employed.