Hemodilutional anemia has been cited as a contributing factor to red blood cell (RBC) transfusions in cardiac surgery patients. Accordingly, efforts have been made to minimize hemodilution by reducing cardiopulmonary bypass (CPB) prime volume. We sought to assess the impact of these efforts on intraoperative RBC transfusions. We evaluated 21,360 patients undergoing coronary artery bypass with or without aortic valve surgery between July 2011 through December 2016 at any of 42 centers participating in the Perfusion Measures and Outcomes registry. The primary exposure was net CPB prime volume (total prime volume minus retrograde autologous prime volume) indexed to body surface area (mL/m2), which was further divided into quartiles (Q1: <262 mL/m2, Q2: 262–377 mL/m2, Q3: 377–516 mL/m2, and Q4: >516 mL/m2). The primary outcome was intraoperative RBC transfusion. We modeled the effect of index net prime volume on transfusion, adjusting for patient (age, gender, race, diabetes, vascular disease, previous myocardial infarction, ejection fraction, creatinine, preoperative hematocrit (HCT), total albumin, status, aspirin, and antiplatelet agents), procedural (procedure types) characteristics, surgical year, and hospital. The median net prime volume was 378 mL/m2 (25th percentile: 262 mL/m2, 75th percentile: 516 mL/m2). Relative to patients in Q1, patients in Q4 were more likely to be older, female, nondiabetic, have higher ejection fraction, have more ultrafiltration volume removed, and undergo more elective and aortic valve procedures (all p < .05). Patients in Q4 relative to Q1 were exposed to lower nadir HCTs on bypass, p < .05. The net prime volume was associated with an increased risk of transfusion (8.9% in Q1 vs. 22.6% in Q4, p < .001). After adjustment, patients in Q4 (relative to Q1) had a 2.9-fold increased odds (ORadj = 2.9, 95% CI [2.4, 3.4]) of intraoperative RBC transfusion. In this large, multicenter experience, patients exposed to larger net prime volumes were associated with greater adjusted odds of receiving intraoperative transfusions. Our findings reinforce the importance of efforts to reduce the net CPB prime volume. Based on these findings and other supporting evidence, the net prime volume should be adopted as a national quality measure.