Background: Blood glucose (BG) measurements are commonly used to screen for neonatal hypoglycemia (NH) among infants admitted to Well Baby Nurseries (WBN). The impact of hypoglycemia screening on WBN care has not been well characterized. Objective: To determine the proportion of initially well-appearing newborns screened for hypoglycemia and describe the indications and outcome of screening over 2.5 years at a single center. Methods: Retrospective cohort study of infants born from 1/1/2013-7/31/2015 at a large maternity center. Study focused on infants who otherwise would receive routine care: birth at ≥36 weeks gestational age (GA); birth weight (BW) ≥2 kg; admitted from delivery suite to WBN. BG values in eligible infants until 72 hours of age were included in the study. Risk criteria for NH screening at our center include: BW < 2500 g or >4000 g; GA 36 0/7-6/7 or ≥41 weeks; BW < 2800 g and gestation >38 weeks; maternal diabetes; in-utero exposure to beta-blockers; symptoms attributable to hypoglycemia. Clinical and laboratory data were obtained from hospital databases and trend analysis performed using linear regression (STATA 14, StataCorp, TX). Results: Study population is shown in Figure 1. Of the 5,140 (48.8%) tested infants, 4,611(89.7%) infants remained in WBN. A total of 16,694 BGs (median BG per infant 3, range 1-24) were obtained from these 4,611 infants in the first 72 hours of life. 2,723/4611 (59.1%) infants had all BG values ≥50 mg/dl and 1,465 (31.8%) had at least one value < 47 mg/dl. Figure 2 depicts the quarterly trends of testing, results and NICU admissions for NH (n = 5,140). The proportion of infants tested (mean 48.9%, SD 2.9) did not change. Among the tested infants, distribution of risk factors (data not shown) and proportion of infants with lowest BG ≤ 35 mg/dl (mean 7.8%, SD 2.4) remained unchanged. However, there was significant increase in the proportion of infants with lowest BG in the 36-46 mg/dl range (mean 26.7%, SD 7.00, p < 0.001); 47-49 mg/dl range (mean 8.6%, SD 2.9, p=0.02); and the proportion of infants admitted to the neonatal intensive care unit (NICU) for hypoglycemia therapy (Mean 4.5%, SD 2.6, p < 0.001) over time. NICU admissions increased from 1.9% to 7.8% between the first and last quarter of the study. Conclusion: Nearly half of all infants initially admitted to WBN met our center criteria for BG testing and one third of those tested met criteria for hypoglycemia. An increasing proportion of tested infants required NICU admission for hypoglycemia despite no apparent change in the distribution of risk factors for hypoglycemia. Critically evaluating the need for such widespread screening, the impact of corrective intervention on long-term outcomes and quantifying the resources used in NH screening can inform clinical and organizational decisions for WBN.