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PBMC cells contain monocytes and lymphocytes. We usually grow the PBMC in TexMACS medium which has been designed for high-performance T cell growth, high cell viability, and consistency under serum-free conditions. Cells are then incubated at 37°C and 5% CO2 for 48 hours. The adherent monocytes are removed and the suspended cells are activated with Anti-Biotin MACSiBead Particles and biotinylated antibodies against human CD2, CD3, and CD28. Anti-Biotin MACSiBead Particles loaded with the biotinylated antibodies are used to mimic antigen-presenting cells and activate resting T cells from PBMCs as well as purified T cells in a bead-to-cell ratio 1:2. Expansion is achieved by adding IL-2 and fresh medium every 3– 4 days. Cells are restimulated at day 14 by adding additional loaded Anti-Biotin MACSiBead Particles at a bead-to-cell ratio of 1:2.