Objective-To determine the efficacy of decontamination and sterilization of a disposable port intended for use during single-incision laparoscopy. Sample-5 material samples obtained from each of 3 laparoscopic surgery ports. Procedures-Ports were assigned to undergo decontamination and ethylene oxide sterilization without bacterial inoculation (negative control port), with bacterial inoculation (Staphylococcus aureus, Escherichia coli, and Mycobacterium fortuitum) and without decontamination and sterilization (positive control port), or with bacterial inoculation followed by decontamination and ethylene oxide sterilization (treated port). Each port underwent testing 5 times; during each time, a sample of the foam portion of each port was obtained and bacteriologic culture testing was performed. Bacteriologic culture scores were determined for each port sample. Results-None of the treated port samples had positive bacteriologic culture results. All 5 positive control port samples had positive bacteriologic culture results. One negative control port sample had positive bacteriologic culture results; a spore-forming Bacillus sp organism was cultured from that port sample, which was thought to be an environmental contaminant. Bacteriologic culture scores for the treated port samples were significantly lower than those for the positive control port samples. Bacteriologic culture scores for the treated port samples were not significantly different from those for negative control port samples. Conclusions and Clinical Relevance-Results of this study indicated standard procedures for decontamination and sterilization of a single-use port intended for use during singleincision laparoscopic surgery were effective for elimination of inoculated bacteria. Reuse of this port may be safe for laparoscopic surgery of animals.