ObjectiveTo compare the humidification efficacy of sterile waterfilled cup and sterile waterfilled water reservoir on infant incubators, as well as  bacterial culture of incubators . MethodsTen infant incubators in a neonatal intensive care unit were randomly divided into observation group and control group, incubators in observation group were humidified through sterile water in single used cups, in control group were through water in water reservoirs , humidified water, air in incubators, and object surface were performed bacterial culture 24 hours after the change of humidified water. ResultsThe humidity in both groups achieved the requirement of >50%; the qualified rate of bacterial culture of humidified water in  observation group was significantly higher than control group(91.00% vs 74.00%,χ2=10.009，P=0.002）;bacterial count in observation group was significantly lower than control group(［26.85±16.67］ CFU/mL vs ［37.20±15.28］CFU/mL，t=4.577，P＜0.001）.The qualified rate of bacterial culture of air and surface of incubators in  observation group were both  significantly higher than control group （94.00% vs 85.00% ;98.00% vs 68.00%，both P＜0.05).ConclusionWaterfilled cups can achieve the air humidification efficacy on infant incubators, reduce the growth of bacteria, and is helpful for preventing the occurrence of healthcareassociated infection.