Background: Early-onset sepsis (EOS) is a potentially fatal condition that affects about 0.3-0.8/1,000 infants born at ≥35 weeks' gestation in developed countries. Current EOS management algorithms result in 8-15% of infants receiving antibiotics for suspected sepsis. The Neonatal Sepsis Calculator provides evidence-based estimates of individual sepsis risk, but data on its clinical application is limited. Objectives: To evaluate the feasibility, safety, and effect on the newborn infants that were investigated and that received antibiotic treatment for suspected EOS following the introduction of the Neonatal Sepsis Calculator. Methods: This was a prospective, observational, single-centre cohort study comparing the rates of newborn infants born at ≥35 weeks' gestation requiring evaluation and/or treatment for suspected EOS in a large tertiary perinatal centre before versus after the prospective introduction of the Neonatal Sepsis Calculator (Epoch 1: October 2014 to January 2015 vs. Epoch 2: July to December 2016). Results: There were 1,732 and 2,502 eligible infants born during Epochs 1 and 2, respectively. Of these, 425 (24.2%) and 530 (21.2%), respectively, were admitted to the neonatal unit. The proportion of infants investigated for sepsis decreased from 15.2 to 11.1%, and that of infants treated with antibiotics from 12.0 to 7.6%. One case of EOS occurred during each Epoch. Conclusions: The implementation of the Neonatal Sepsis Calculator was feasible and safe in our unit. Application of this clinical decision support tool may reduce the number of infants undergoing investigations and empirical treatment for suspected EOS.