Objectives To evaluate whether implementing more restrictive neonatal intensive care unit (NICU) platelet transfusion guidelines following the Platelets for Neonatal Transfusion - Study 2 randomized controlled trial (transfusion threshold changed from 50 000/μL to 25 000/μL for most neonates) was associated with fewer NICU patients receiving a platelet transfusion, without adversely affecting outcomes. Study Design Multi-NICU retrospective analysis of platelet transfusions, patient characteristics, and outcomes during 3 years before vs 3 years after revising system-wide guidelines. Results During the first period, 130 neonates received 1 or more platelet transfusions; this fell to 106 during the second. The transfusion rate was 15.9/1000 NICU admissions in the first period vs 12.9 in the second (P = .106). During the second period, a smaller proportion of transfusions was administered when the platelet count was in the 50 000–100 000/μL range (P = .017), and a larger proportion when it was <25 000/μL (P = .083). We also saw a fall in the platelet counts that preceded the order for transfusion from 43 100/μL to 38 000/μL (P = .044). The incidence of adverse outcomes did not change. Conclusions Changing platelet transfusion guidelines in a multi-NICU network to a more restrictive practice was not associated with a significant reduction in number of neonates receiving a platelet transfusion. The guideline implementation was associated with a reduction in the mean platelet count triggering a transfusion. We speculate that further reductions in platelet transfusions can safely occur with additional education and accountability tracking. To evaluate whether implementing more restrictive neonatal intensive care unit (NICU) platelet transfusion guidelines following the Platelets for Neonatal Transfusion - Study 2 randomized controlled trial (transfusion threshold changed from 50 000/μL to 25 000/μL for most neonates) was associated with fewer NICU patients receiving a platelet transfusion, without adversely affecting outcomes. Multi-NICU retrospective analysis of platelet transfusions, patient characteristics, and outcomes during 3 years before vs 3 years after revising system-wide guidelines. During the first period, 130 neonates received 1 or more platelet transfusions; this fell to 106 during the second. The transfusion rate was 15.9/1000 NICU admissions in the first period vs 12.9 in the second (P = .106). During the second period, a smaller proportion of transfusions was administered when the platelet count was in the 50 000–100 000/μL range (P = .017), and a larger proportion when it was <25 000/μL (P = .083). We also saw a fall in the platelet counts that preceded the order for transfusion from 43 100/μL to 38 000/μL (P = .044). The incidence of adverse outcomes did not change. Changing platelet transfusion guidelines in a multi-NICU network to a more restrictive practice was not associated with a significant reduction in number of neonates receiving a platelet transfusion. The guideline implementation was associated with a reduction in the mean platelet count triggering a transfusion. We speculate that further reductions in platelet transfusions can safely occur with additional education and accountability tracking.