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Objective: To evaluate the disinfection effect of high-energy pulse ultraviolet disinfection equipment in medical institution settings. Methods: The disinfection effect was evaluated through field tests and laboratory tests. Among them, 135 high-frequency contact points were selected from nine departments in the field test. Samples were collected before and after disinfection, and the disinfection effects of 75% alcohol wipes wiping disinfection, high-energy pulse ultraviolet disinfection robot disinfection and high-energy pulse ultraviolet handheld disinfection instrument were compared. In the laboratory test, 30 infected areas of the simulated test table were exposed to vertical ultraviolet irradiation and the bacterial-killing rate before and after disinfection was calculated. Results: In the field test, the bacteria-killing rates of 75% alcohol wipes, high-energy pulse ultraviolet disinfection robot and high-energy pulse ultraviolet handheld disinfection instrument were 94.99%, 91.53% and 95.94%, respectively, and the difference was statistically significant. The disinfection effect of the high-energy pulse ultraviolet handheld disinfection instrument was better than that of the high-energy pulse ultraviolet disinfection robot ( P values <0.05). In the laboratory test, the killing log value of Staphylococcus aureus and Escherichia coli on the carrier were both greater than 3.00. In the simulated field test, the killing log value of Staphylococcus aureus on the surface samples were 4.99. Conclusion: Both the high-energy pulse ultraviolet handheld disinfection instrument and the high-energy pulse ultraviolet disinfection robot have good disinfection effects, which are similar to the disinfection effects of conventional 75% alcohol wipes.