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Introduction: Reduction in innate immune function is associated with adverse outcomes in critically ill adults and children. Our prior work suggests that RBCs suppress monocyte function in vitro as a function of storage age. However, in vivo effects of a single RBC transfusion in critically ill children are unknown. Hypothesis: In critically ill children, a single transfusion of RBC with longer (vs. shorter) storage time will be associated with reduced immune function over time (decreased ability of whole blood to produce TNFα upon ex vivo stimulation with LPS). Methods: This is an ongoing multi-center prospective observational study in which children in one of two quaternary-care pediatric ICUs with an order for RBC transfusion are included. Patients with emergent transfusion, severe leukopenia, recent blood product transfusion (within one month), or history of transplant are excluded. Blood is drawn immediately pre-transfusion and again 18-30 hours post-transfusion. 50µl of whole blood is added to 500µl of stimulation solution containing 500pg/ml LPS and incubated for 4hrs at 37C. TNFα is measured in the supernatant by chemiluminescence. Data represent median [IQR]. Results: 22 subjects have been enrolled to date (age: 52 [4 – 99] months, time from ICU admission to transfusion: 2 [0.9 – 4.2] days, transfused volume: 15 [12-18] ml/kg). Children whose RBC were stored >21 days (n=7, median storage time: 29d) showed worsening of immune function compared to children whose RBC storage times were shorter (median storage time: 7d): TNFα production capacity 94% vs 150% of pre-transfusion value (p=0.03, Mann Whitney). Shorter RBC storage time was associated with improvement in TNFα production capacity over time (odds ratio: 8.7 [1.1-72]). Patient age, time from ICU admission to transfusion, and PRISM III score at the time of transfusion were not significantly different between groups. Lower thresholds of RBC storage time (7, 14 days) were not associated with immune impairment in this cohort. Conclusions: A single transfusion of RBC stored for > 21 days is associated with reduced immune function in critically ill children. Further studies are warranted to determine clinical impact and mechanisms of RBC-induced immunosuppression.