Fig. 3. Analysis of GFP⁺ and GFP^– EB-derived cells. (A) At day 3.5 of differentiation, GFP Bry EBs were harvested, dissociated and cells fractionated by cell sorting based on their GFP expression level. (B) RNA extraction was performed on the GFP^–, GFP⁺ as well as the starting population (presort). Gene expression patterns were analyzed by RT-PCR using specific primers for each of the indicated genes. (C) Blast-CFC potential of the presort, GFP^– and GFP⁺ populations. Colonies were scored after 4 days of culture. Data are presented as the mean number of colonies from three dishes. Bars, where visible, represent s.e. of the mean. The purity of sorted cells was 95%. (D) Expression analysis of day 6 reaggregated EBs generated from day 2.5 EB presort, GFP⁺ and GFP^– cells. (E) Neurite potential of reaggregated EBs generated from GFP⁺ and GFP^– populations. Data are presented as the % of EBs that form neurites. (F) Immunostaining demonstrating the expression of neuronal class III ß-tubulin (TuJ1) on neurites that develop from EBs derived from GFP^– cells.

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