A fully automated method for mononuclear bone marrow cell concentration

Abstract

We describe our experience in processing 40 bone marrow aspirates harvested for autotransplantation from patients with several hematological diseases using the CS-3000 blood cell separator. The bone marrow of the first 30 patients was processed by a semiautomated method, and a fully automated procedure was used for the remaining 10 cases. Both procedures were developed in our laboratory and yielded a similar average mononuclear cell recovery of 87.78% and 86.98%, respectively, and similar nucleated cell recovery (27.39% and 27.11%). The cloning efficiency of hematopoietic progenitor cells, measured as the total CFU-GM colony recovery in the in vitro cultures, did not differ between processed and recovered mononuclear cells. On the other hand, all the patients with transplants showed complete hematologic recovery, and the time to engraftment was similar to that described for other procedures. The automated procedure resulted in an average red cell removal of 97.81%, similar to the semiautomated procedure (94.19%), though with a narrower range (96.31-98.6% vs. 80.34-98.34%). The time taken to process a similar amount of bone marrow cell suspension was very different for each method: 1 hour for the fully automated vs. 2 1/2 hours for the semiautomated method to process 1,000 ml. Furthermore, the semiautomated procedure required the addition of homologous or irradiated plasma in a laminar air flow chamber, while the automated method is performed in a closed sterile system. We conclude that our procedure using the CS-3000 processor is an efficient method for fully automated large-scale processing of human bone marrow cells.

Hematopoietic stem cell transplantation between red cell incompatible donor–recipient pairs

“…7,[27][28][29] The sedimentation of red cells can be enhanced by the addition of macromolecules such as hydroxyethylstarch to the component. 30 Any of the apheresis devices can be used to separate red blood cells from the bone marrow component.…”

“…30 Any of the apheresis devices can be used to separate red blood cells from the bone marrow component. 7,[27][28][29] The processed component usually contains 5-20 ml of red blood cells. The recov- In situations where cell dose may be important to transplant outcome, 31 the minimal cell losses of red cell depletion can be avoided by isoagglutinin reduction of the recipient.…”

Analysis for Recovery and Loss of Mononuclear Cells and Colony-Forming Units Granulocyte-Macrophage during ex vivo Processing of Autologous Bone Marrow

“…Reports using the Cobe system with or with out density separation media resulted in mean recovery rates of 58-85% for MNC and greater than 80% for CFU-GM [6,8,24,26,27]. BM processing with the Fenwal CS 3000 apheresis system resulted in mean recoveries of 53-86% for MNC and 76-98% for CFU-GM [9,10].…”

“…Contaminating granulocytes can in terfere with the activity of monoclonal antibodies and com plement during purging procedures [18]. After thawing ly sis of granulocytes may lead to clumping of the marrow by release of lysosomal enzymes and high amounts of lysed RBC may cause postinfusion hemoglobinuria [8,10]. Therefore, the BM processing technique should generate a low volume concentrate with a maximum recovery of MNC and CFU-GM and a minimum RBC and granulocyte con tamination.…”

“…As the hematopoietic stem and pro genitor cells are found in the MNC population, a mononu clear cell concentrate is usually prepared for cryopreservation using semiautomated blood cell processors with or without density gradients like Ficoll or Percoli [6][7][8]. Re cently, large-scale processing of human BM with automated blood cell separators have been described suggesting im provements in terms of cell recovery, collection selectivity and microbiologie safety in a closed system [9][10][11]. We in vestigated the recovery of MNC and CFU-GM in 25 autolo gous BM concentrates prepared for cryopreservation with the Fresenius AS104 blood cell separator.…”