To evaluate the impact of the implementation of universal bilirubin screening on neonatal health care use in the context of a large jurisdiction with universal health insurance.We conducted a population-based retrospective cohort study of all newborns discharged after birth between April 2003 and February 2011 from 42 hospitals that implemented universal bilirubin screening between July 2007 and June 2010 in Ontario, Canada. We surveyed hospitals to determine their screening implementation date. We used multiple linked administrative health data sets to measure phototherapy use, length of stay (LOS), jaundice-related emergency department visits, and jaundice-related readmissions. We modeled the relationship between universal bilirubin screening and outcomes using generalized estimating equations to account for clustering by hospital, underlying temporal trends, and important covariates.Screening was associated with an increase in phototherapy during hospitalization at birth (relative risk, 1.32; 95% confidence interval, 1.09-1.59) and a decrease in jaundice-related emergency department visits (relative risk, 0.79; 95% confidence interval, 0.64-0.96) but no statistically significant difference in phototherapy after discharge, LOS, or jaundice-related readmissions after accounting for preexisting temporal trends in health care service use and other patient sociodemographic and hospital characteristics.Universal bilirubin screening may not be associated with increased neonatal LOS or increased subsequent hospital use. Our findings are relevant for determining the resource implications of universal bilirubin screening in Ontario. They highlight the limitations in generalizability of previous research on health care utilization associated with similar programs and underline the importance of context-specific local evaluation of guideline implementation.