Background: Neonatal thrombosis is a frequently encountered complication in a neonatal intensive care unit. Dalteparin can be used to treat thrombosis in newborn infants. Objectives: In this study, we evaluate the current recommended starting dose of 129 ± 43 U/kg/24 h, hypothesizing that this dose is too low to reach therapeutic anti-Xa levels. Methods: From 2008 until 2017, all infants treated with dalteparin in the University Medical Centre Utrecht were included in this study. In this retrospective cohort study, the correlation between dose and anti-Xa level was observed. Results: Sixty-six infants were included. The most common thrombus types were catheter-related (29 patients, 44%) and venous sinus thrombosis (28 patients, 43%). The mean dalteparin dose needed for the first adequate anti-Xa level (0.5–1.0 IU/mL) was 297.6 U/kg/12 h. Two infants developed a first bleeding episode under dalteparin therapy; they both had anti-Xa levels in the therapeutic range. Conclusion: The increase of the starting dose of dalteparin will lead to earlier therapeutic levels of anti-Xa in the studied population and appears to be safe. However, this needs to be evaluated in further study.