Efficient elutriation of monocytes within a closed system (Elutra™) for clinical-scale generation of dendritic cells

Dendritic cells (DC) are promising tools for the immunotherapy of cancer. The induction of tumor-specific T cells and clinical regressions have already been observed in early phase I/II vaccination trials. As DC vaccination is now facing trials with larger patient collectives it becomes increasingly important to obtain large numbers of cells suitable for therapeutic applications under labor- and cost-effective conditions. We describe here a procedure that uses a novel cell separator (Elutra (TM), Gambro BCT) to enrich monocytes from an entire apheresis product within one hour. Cells are separated on the basis of size and to a lesser extent density, by elutriation in a 40-ml conical chamber. The total monocyte recovery following elutriation (n=6) was 98.53% (+/- 8.07%), the recovery in the monocyte-rich fraction 75.45% (+/- 11.31%), and the mean purity 82.95% (+/- 6.01%). These monocytes can be cultured either in conventional culture dishes or in closed cell culture bags and differentiated, by using GM-CSF+IL-4 followed by a maturation cocktail composed of IL-1 beta+IL-6+TNF-alpha+PGE(2), into fully mature DC. The Elutra (TM) separator allows for fast and easy enrichment of monocytes within a closed system. Subsequently, elutriated monocytes can be successfully cultured into phenotypically and functionally mature DC for immunotherapeutic approaches. The method neither requires a density gradient step to enrich PBMC from leucapheresis products nor does it apply (xenogeneic) antibodies to target monocytes. Isolation of monocytes with Elutra (TM) may greatly facilitate future DC-based vaccination approaches. (c) 2005 Elsevier B.V. All rights reserved.