The method described here using a centrifuge and Coulter Cell Counter for the quantitative differential agglutination of human red cells uses commercial anti-A and anti-B antisera for the ABO system, and for the Rh system a commercial anti-D serum, a low ionic strength solution and an anti-human IgG antiserum. We compared this Coulter Counter method with the Technicon Auto Analyzer method which utilizes bromelin and polyvinyl pyrrolidone, and anti-A, anti-B and anti-CD antisera, and found this new method to be the simpler of the two. The nonagglutinable count with the Coulter Counter was 1.07% for A(1) red cells, 2.26% for A(2) red cells, 1.06% for B red cells, and 1.78% for Rh-positive red cells, results similar to those seen with the Technicon Auto Analyzer. Results with the Coulter Counter method were consistently accurate whether the ACD red cells were studied on the day of collection, after 10 days of 4 °C storage, or after 4 °C storage for up to 6 days followed by cryopreservation with 40% (w/v) glycerol at -80 °C, thawing and washing. In this study, red cell samples obtained from recipients who had received compatible but identifiable donor red cells were frozen with 40% glycerol and stored at -80 °C for 10 months, thawed and washed. Survival measurements on these washed previously frozen red cells were similar to the values in liquid-stored red cells.
Baboon red cells were treated to reduce oxygen affinity by an osmotic-pulse procedure using dimethylsulfoxide. Inositol hexaphosphate (IHP) alone or IHP and adenosine triphosphate (ATP) were incorporated into the red cells in the presence of polyethylene glycol (PEG). The procedure produced variable increases in the red cell P(50) value, i.e., the partial pressure of oxygen at which 50% of the hemoglobin was saturated. The effect of treatment of autologous baboon red cells on the 24-hour posttransfusion survival value and lifespan T(50) value was measured using a double-label procedure. The data demonstrate that the increase in the P(50) value of treated red cells was negativelycorrelated with the 24-hour posttransfusion survival value; the higher the P(50) value, the poorer the 24-hour posttransfusion survival value. The 24-hour posttransfusion survival value for nontreated baboon red cells was 90% and the T(50) value was 14 days. The IHP-ATP-PEG-treated red cells had significantly higher red cell ATP levels than did IHP-PEG-treated red cells. The 24-hour posttransfusion survival value was 68% for the IHP-ATP-PEG treated red cells and 52% for the IHP-PEG-treated red cells when the increase in P(50) ranged from 10 to 20 mm Hg; the lifespan T(50) value for both the IHP-ATP-PEG-treated red cells and the IHP-PEG-treated red cells was 15 days. Osmotic pulse treatment produced significant red cell injury manifested by the 24-hour posttransfusion survival value. However, modification of the RBC with IHP-PEG or IHP-ATP-PEG to decrease hemoglobin affinity for oxygen did not affect their lifespan.
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