Hematopoietic Cell Colony-Forming Assay

Briefly, a single cell suspension of mononuclear cells from mouse bone marrow was prepared to obtain ~2–4 × 10⁷ hematopoietic cells using previous described methods (26 (link)). The cells were resuspended in 10mL of Iscove’s modified Dulbecco’s medium (IMDM/2% FBS media) (Allcells) and mixed with Methyl cellulose (R&D systems), and 1.1mL of the final cell mixture was added to a non-treated 6-well plate using a 3mL syringe. Water was added to one of the wells of the 6-well plate to maintain humidity necessary for colony development. Plates were incubated for 8–12 days at 37°C and 5% CO₂. Colonies consisting of at least 30 cells were counted.