Figure 5.

Antigen-uptake capacity of dendritic cells (DC), generated by culture with granulocyte–macrophage colony-stimulating factor (GM-CSF), with or without interleukin-10 (IL-10). DC were cultured with GM-CSF, with or without IL-10. On day 9, DC were incubated with dextran–fluorescein isothiocyanate (FITC) for 1 hr at 37°, or at 0° as a control. Afterwards, the cells were washed twice with ice-cold medium. (a) Cells were stained for major histocompatibility complex (MHC) class II expression and analysed on a flow cytometer, using 7-aminoactinomycin D (7-AAD) to exclude dead cells. (b) Expression of CD40, CD80 and CD86 on gated MHC class II^dim dextran–FITC^negative cells (thin line) and on gated MHC class II^dim dextran–FITC^positive cells (thick line) was analysed by four-colour flow cytometry (dextran–FITC/CD40^– CD80^– or CD86–phycoerythrin (PE)/7-AAD/MHC class II biotin + streptavidin-allophycocyanin (APC)]. (c) MHC class II^dim dextran–FITC^negative cells and MHC class II^dim dextran–FITC^positive cells were sorted using a FACScalibur. Graded numbers of both sorted and the presort DC were cultured with purified allogeneic CD4⁺ T cells. Proliferation was assessed after 4 days by measuring the incorporation of [³H]thymidine.