Optimization of biotinylation protocol for next generation studies of red blood cell survival after transfusion

Red blood cell (RBC) biotinylation has increasingly become the preferred method for quantifying RBC post-transfusion recovery (PTR). This study evaluated the feasibility of transfusing autologous RBCs labeled with biotin (BioRBCs) 48 h prior to transfusion. The rationale was to facilitate the distribution of BioRBC products from manufacturing sites to remote clinical sites.

Leukocyte-reduced RBC units from 12 healthy individuals were stored at 1-6°C for 42 days and biotinylated with two biotin densities (3 and 15 μg/mL) 48 and 4 h prior to transfusion. Each individual was transfused with 10 mL of each BioRBC preparation, after which BioRBC 20 h PTR and long-term survival (30 and 90 days) were determined by flow cytometric analyses. Additional quality measurements included phosphatidylserine (PS) exposure on BioRBCs and in vitro metrics of hemolysis.

RBC transfusion 48 h after biotinylation was not associated with altered BioRBC 20 h PTR or 30 and 90 day survival as compared with RBC transfusion 4 h after biotinylation (e.g., PTR 20 h: 85.7% ± 8.4% vs. 87.5% ± 7.3%; 48 and 4 h, respectively, p > .05). Similarly, no significant differences between the BioRBC groups were observed in BioRBC PS exposure at all time points. BioRBC long-term survival, but not 20 h PTR, was negatively associated with donor hemoglobin (Pearson r = -0.71, p = .001) and osmotic hemolysis (r = -0.783, p = .003). None of the participants developed antibodies against BioRBCs during the trial.

RBC biotinylation 48 h prior to transfusion does not compromise the quality and safety of BioRBC products. BioRBC long-term survival can be used to identify donor characteristics that influence RBC lifespan in the circulation.