Stem cell factor (scf)

Frozen human UCB CD34⁺ HSPCs were purchased from StemCell Technologies (Vancouver, BC, Canada) and used as provided (passage number 1). The CD34⁺ cell purity in the HSPCs was determined to be 98% by flow cytometry, and the viability was determined to be >97% by trypan blue staining.
CD34⁺ HSPCs were cultured in StemSpan SFEM II expansion medium (StemCell Technologies) with 100 ng/ml SCF (ProSpec, Rehovot, Israel), 100 ng/ml Flt-3L (ProSpec), 50 ng/ml TPO (ProSpec), 20 ng/ml IL-3 (ProSpec), and 20 ng/ml IL-6 (ProSpec). The initial inoculum and maintenance densities were 5 × 10⁴ and 1 × 10⁶ cells/ml, respectively. Culture media were exchanged with fresh media at one-half volume of the initial media every 3 days, and the cultures were maintained at 37°C in a humidified atmosphere containing 5% CO₂.

Primary Ph⁺ ALL bone marrow cells were kindly provided by Dr Michael Caligiuri (Ohio State University, Columbus, OH) and Dr Martin Carroll (University of Pennsylvania, Philadelphia, PA). Cells were seeded on a substrate of adherent, mitomycin C-treated OP9 stromal cells and maintained in SFEM (Stem Cell Technology, Vancouver, Canada) supplemented with IL-3 (10 ng/ml), IL-7 (10 ng/ml), FLT3L (20 ng/ml) and SCF (30 ng/ml) (ProSpec, Israel).
CD34⁺ CML cells were kindly provided by Dr Tessa Holyoake (University of Glasgow, United Kingdom) and cultured in SFEM supplemented with IL-3 (20 ng/ml), IL-6 (20 ng/ml), SCF, and thrombopoietin (10 ng/ml).
Commercially purchased (Stem Cell Technologies) cord blood CD34⁺ cells were cultured in SFEM (Stem Cell Technologies) enriched with the CC100 cytokine cocktail (SCF, 100 ng/ml; FLT3L, 100 ng/ml; IL-3, 20 ng/ml; IL-6, 20 ng/ml).
Ph⁺ and normal CD34⁺ primary cells were kept in culture at 37 °C, under a 5% CO₂ humidified atmosphere. Cell counts were performed using 0.4% Trypan Blue Solution and a Neubauer hemocytometer.