A new technique for peripheral stem cell separation

Using a Haemonetics V50.1 cell separator we compared the new lymphocyte-surge-program for peripheral stem cell preparation with the conventional combined buffy coat and Ficoll techniques. Eleven surge and four conventional separations were performed in 15 healthy cytapheresis donors who gave informed consent to the procedure. Using the conventional methods, mean recovery of peripheral blood mononuclear cells (PBM) was 55% in a preparation time of 80 minutes for the buffy coat and 25 minutes for the Ficoll step. Eighty percent of the PBM loss occurred during the buffy coat step, and 20% during the Ficoll procedure. In a preparation time of 105 minutes, 2500 mL peripheral blood could be processed without flow or citrate problems. The donors' red cell losses were about 100 mL packed red cells. Granulocyte depletion was about 98%, red cell depletion higher than 99% (hematocrit < 0.1%). Using the surge technique we achieved a PBM recovery of 60% (range 45% to 72%) by processing 2100 mL peripheral blood in a preparation time of only 75 minutes. The donors' red cell losses were about 8 mL of packed red cells, significantly lower (P < 0.01) compared to conventional techniques. Nevertheless, red cell contamination of the concentrates remained high (hematocrit = 1.8%). Colony-forming unit (CFU) tests showed comparable quantities of peripheral stem cells (90 colonies/10⁷ PBM) in both separation techniques. Therefore, both techniques seem to be adequate for T-cell depletion and preparation of bone marrow for cryopreservation. Since preparing bone marrow for purging techniques requires extreme stem cell purity, the surge technique may be less suitable for those purposes due to the awkward problem of considerably higher red cell contamination, despite a shorter preparation time.