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To obtain T cells for CAR-T cell production, blood is collected and leukapheresis is performed to isolate and extract peripheral mononuclear blood cells (PBMCs, e.g., leukocytes and lymphocytes). The T cells in this PMBC fraction are then separated out using cell processors or centrifugation-based cell separators. Cell surface marker-mediated techniques (e.g., magnetic-activated cell sorting, MACS) can be employed to further isolate specific T cell subsets.1
Sustained T cell activation is needed for ex vivo expansion. This can be accomplished by co-incubating T cells with natural or artificial antigen-presenting cells (APCs). Alternatively, antibody-coated beads can deliver the necessary signals to the T cells, and they have become popular to both select for and activate T cells in a single step.1
1. X. Wang and I. Rivière, “Clinical manufacturing of CAR T cells: foundation of a promising therapy,” Mol Ther Oncolytics 3:16015, 2016.
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