Banked blood exhibits impairments in nitric oxide (NO)-based oxygen delivery capability, reflected in rapid depletion of S-nitrosohemoglobin (SNO-Hb). We hypothesized that transfusion of even freshly-stored blood used in pediatric heart surgery would reduce SNO-Hb levels and worsen outcome. In a retrospective review (n = 29), the percent of estimated blood volume (% eBV) replaced by transfusion directly correlated with ventilator time and inversely correlated with kidney function; similar results were obtained in a prospective arm (n = 20). In addition, an inverse association was identified between SNO-Hb and postoperative increase in Hb (∆Hb), reflecting the amount of blood retained by the patient. Both SNO-Hb and ∆Hb correlated with the probability of kidney dysfunction and oxygenation-related complications. Further, regression analysis identified SNO-Hb as an inverse predictor of outcome. The findings suggest that SNO-Hb and ∆Hb are prognostic biomarkers following pediatric cardiopulmonary bypass, and that maintenance of red blood cell-derived NO bioactivity might confer therapeutic benefit.