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Whether critically ill neonates needing a surgical intervention should be transferred to an operating room (OR) or receive the intervention in a neonatal intensive care unit (NICU) is controversial. In this study, we report our experience in performing surgical procedures in a NICU including air cleanliness.This was a retrospective study performed at a metropolitan hospital. The charts of all neonates undergoing surgical procedures in the NICU and OR were retrospectively reviewed from January 2007 to June 2017. Data on baseline characteristics, procedure and duration of surgery, ventilator use, hypothermia, instrument dislocations, surgery-related infections and complications, and outcomes were analyzed.Ninety-two neonates were enrolled in this study, including 44 in the NICU group and 48 in the OR group. The air cleanliness was International Organization for Standardization (ISO) 14644-1 class 7 in the NICU and class 5-6 in the OR. The NICU group had a younger gestational age and lower birth body weight than the OR group. The OR group had a higher incidence of hypothermia than in the NICU group (56.3% vs 9.1%, P < .001). However, there were no significant differences in surgical site related infections or mortality between the 2 groups.This study suggests that performing surgical procedures in a NICU with air cleanliness class 7 is as safe as in an OR, as least in part, when performing patent ductus arteriosus ligation and exploratory laparotomy.