Indirect induction of DCs from p14⁻ cells via a CD14⁺ intermediate precursor. (A) p14⁻ cells sorted from a 28-day primary culture with FLT3-L plus TPO and SCF were incubated with GM-CSF plus FLT3-L for 3 and 7 days. CD14⁺ cells generated from the 3-day culture in GM-CSF plus FLT3-L were sorted out and all cell suspensions were additionally cultured for 4 days in GM-CSF plus IL-4 (bottom histograms). (B) Alloreactivity of some of the cellular fractions depicted in (A). GF-GF: p14⁻ cells incubated for 7 days in GM-CSF plus FLT3-L, corresponding to the upper right histogram in (A); GF-G4: p14⁻ fraction incubated 3 days in GM-CSF and FLT3-L, then 4 days with GM-CSF and IL-4, corresponding to the lower left histogram in (A); GF-G4-GT4: p14⁻ fraction incubated 3 days in GM-CSF plus FLT3-L, 4 days with GM-CSF and IL-4, and 4 more days with these cytokines plus TNF. Counts per minute are displayed in a log scale to facilitate end point reading of low values. The last significant value over background is determined as in Fig 4A. Mean ± SD of quadruplicates are shown.