Purified primary CD34+ cells were cultured in Stemspan serum-free medium (Stem Cell Technologies) containing 10 ng/mL human SCF, and 10 ng/mL human TPO (Peprotech) in the presence of metformin. Forty-eight hours after the culture, the cells were transferred to a methylcellulose-based medium (Methocult H4034 Optimum, StemCell Technologies) in triplicate, and colonies were manually counted after 10–14 days.
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