Functionality of cRBC hemoglobin. (A) Hemoglobin status of cRBCs determined by HPLC (Bio-Rad Variant II). The percentage of hemoglobin in the elution peak is indicated for the Hb0, HbF, HbA1c, and HbA2 fractions. One representative graph from 9 independent experiments is shown. (B) Tonometric oxygen binding curves at 37°C for a cRBC (triangles) and a control RBC suspension (dotted line) at different DPG/Hb₄ ratios in 10mM HEPES buffer (pH 7.4) containing 150mM NaCl. The RBC isotherms were simulated from the average MWC parameters for 10 different blood samples (black line).