EHZF expression in CD34⁺ cells. (A) Flow cytometric analysis of purified CD34⁺ cells tested with anti–CD34-FITC and anti–CD38-PE. (B) Flow cytometric analysis of purified CD34⁺ cells after 7 days of cytokine treatment (GM-CSF, IL3, and Epo) with anti–CD14-FITC and anti–CD34-PE antibodies. (C) CD34⁺ cells were treated with cytokines for 1, 2, 5, and 7 days and tested with anti–CD14-FITC and anti–CD34-PE antibodies. The percentage of cells positive for CD14 (▪) and for CD34 (□) is shown. (D) Total RNA was prepared and analyzed by Northern blotting from peripheral blood leukocytes (10 μg), purified CD34⁺ cells either untreated (5 μg) or exposed to GM-CSF, IL3, and Epo for 1, 2, 5, or 7 days (5 μg, total). The Northern blot was first hybridized with an EHZF probe, washed and exposed for 7 days, and subsequently rehybridized with a GAPDH probe, washed and exposed for 20 hours. The down-regulation of EHZF was apparent in 2 additional CD34⁺ differentiation time courses analyzed by semiquantitative PCR (not shown).