Immunophenotypic and morphologic analyses of DC-like cells.

13dpc FL-derived Lin⁻c-kit⁺ HPCs were first supplemented with PA6 cells + GM-CSF + SCF + Flt3L for 12 to 14 days and then stimulated with GM-SCF + mTNFα for an additional 3 to 5 days. (A) The phenotype of the nonadherent cells was analyzed by 2-color immunofluorescence staining as described in the Materials and Methods. The indicated FITC-labeled MoAbs (CD8α, NK1.1, F4/80, CD40, CD11c, DEC205, and E-cadherin) were used to demonstrate the phenotypic characteristics of the generated DCs. The quads were set up on the isotype-matched control dot plot. (B) Giemsa staining and nonspecific esterase activity analyses were performed on GM-CSF + mTNFα-stimulated DC precursors at day 3 to day 5 that were derived from FL-derived HPCs cocultured with PA6 cells + GM-CSF + SCF + Flt3L for 12 to 14 days. Original magnification: × 400. These results are representative of 3 independent experiments.