Biotinylation of Adult Donor Allogeneic RBCs for Neonatal Transfusion

Adult donor allogeneic RBCs administered to neonates were biotinylated as previously described.^{18 (link)} Briefly, RBCs were washed using a carbonate-buffered dextrose sodium phosphate wash solution to remove plasma and resuspended at a hematocrit of 25%. Hematocrit adjustment was verified using the Sysmex XE-2100 (Sysmex, Kobe, Japan). Subsequently, RBCs were biotinylated using the biotin 3-sulfo-N-hydroxysuccinimide ester (EZ-LINK Sulfo-NHS-Biotin; Pierce Chemical, Rockford, IL 61101). Depending on the primary objectives of the original study, up to four separate concentrations ranging from 6 to 162 μg of sNHS-biotin per mL of packed RBCs were chosen. These choices produce discreet biotin labeling densities that can be individually tracked for at least 95% of the RBC lifetime. Minutes prior to transfusion, a 0.5 mL sample of the BioRBC mixture was taken for bacterial culture.