OBJECTIVE: To estimate the cost and cost-effectiveness of universal newborn hearing screening. STUDY DESIGN AND SETTING: Decision analysis model utilizing the hospital perspective. This model evaluated 4 distinct protocols for screening a fixed and defined hypothetical cohort of newborn infants. OUTCOME MEASURES: Cost of screening and the number of infants with hearing loss identified through universal screening. RESULTS: Otoacoustic emissions testing at birth followed by repeat testing at follow up demonstrated the lowest cost ($13 per infant) and had the lowest cost-effectiveness ratio ($5100 per infant with hearing loss identified). Screening auditory brainstem evoked response testing at birth with no screening test at follow-up was the only protocol with greater effectiveness, but it also demonstrated the highest cost ($25 per infant) and highest cost-effectiveness ratio ($9500 per infant with hearing loss identified). These findings were robust to sensitivity analysis, including best-case and worst-case estimation. The prevalence of hearing loss and the fraction of infants returned for follow-up testing had a large impact on the absolute level, but not relative level of protocol cost and cost-effectiveness. CONCLUSION: The otoacoustic emissions testing protocol should be selected by screening programs concerned with cost and cost-effectiveness, although there are certain caveats to consider. SIGNIFICANCE: The most significant barriers to implementation of universal newborn hearing screening programs have been financial, and this study compares the most common protocols currently in use. This study can assist program directors not only in the decision to initiate universal screening but also in their choice of screening protocol.