Human peripheral blood was collected from healthy donors. PBMCs were isolated using density-gradient centrifugation with Ficoll-Paque PLUS (GE Healthcare, Milwaukee, WI, USA) for 20 min at 1000 g. The PBMC fractions were collected and then transferred to a new 15 ml centrifugation tube. The PBMCs were washed twice with RPMI 1640. Finally, the PBMCs were resuspended at an appropriate density in RPMI 1640 supplemented with 10% FBS for further experiments.
Bone marrow cells were isolated from C57BL/6 mice and cultured in the presence of 10 ng/ml recombinant murine GM-CSF and IL-4 (Pepro Tech, London, UK). BMDCs from 5-day-old cultures were collected and/or enriched with CD11c MicroBeads (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany) for further experiments.
Bone marrow cells were isolated from C57BL/6 mice and cultured in the presence of 10 ng/ml recombinant murine GM-CSF and IL-4 (Pepro Tech, London, UK). BMDCs from 5-day-old cultures were collected and/or enriched with CD11c MicroBeads (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany) for further experiments.