Fig. 3. Dll1 overexpression in a small proportion of NPCs induces neuronal differentiation in vitro. Primary NPCs were prepared from (A-D) 3-day sphere cultures [3 days in vitro (3 DIV)] or (E,F) 12-day sphere cultures (fourth passage, 12 DIV) of the neocortical cells of E12.5 ICR mice. The NPCs were infected with a retrovirus encoding GFP (pMX-GFP, control), or both GFP and Dll1 (pMX-Dll1-IG), at a low titer and subjected to clonal analysis (see Materials and methods). (A) A representative GFP-positive clone stained with anti-GFP, TuJ1 and Hoechst. After incubation for 2 days in the presence of a low dose of human FGF2 (2 ng/ml) (B-D) or in the absence of FGF2 (E,F), cells were stained with anti-GFP and TuJ1 or anti-GFP and anti-Gfap. The percentage of clones containing only TuJ1-positive (B,E) or only Gfap-positive (D,F) cells among GFP-positive clones was then determined by immunocytochemical analysis. The percentage of TuJ1-positive cells among GFP-positive cells was also determined by immunocytochemistry (C). ^*P<0.02, ^**P<0.01, ^***P<0.0001. Scale bar: 50 µm in A.