The ζ to α switch occurs during terminal erythroid maturation. (A) Two possible models to explain the switch observed in liquid culture. (B) Histograms illustrating globin expression determined by real-time PCR on 14 clonal populations obtained by liquid culture of sorted individual CD34⁺ cells produced in 2-week cocultures of hESCs with FH-B-hTERT. (C) Morphology of cells obtained after 14 or 24 days of liquid culture of CD34⁺ cells from a 20-week-old fetal liver. After 14 days, most cells are pro- or basophilic erythroblasts. After 24 days, most cells are orthochromatic or enucleated red cells. (D) Chromatograms illustrating globin expression in the cells depicted in panel C. The pro- and basophilic erythroblasts express ζ-globin in much larger amounts than the orthochromatic and enucleated red cells. (E) Histograms summarizing the quantification of the results illustrated in panel D. (F) Histograms summarizing a real-time PCR analysis of globin expression performed on the cells described in panel C. The results suggest that at least part of the regulation occurs at the transcriptional level. Percent α-globin was calculated as 100 × α-globin/(α-globin + ζ-globin). Percent γ-globin was calculated as 100 × γ-globin/(γ-globin + ϵ-globin). Error bars represent SD.