CD48^−/− HSCs are less proliferative in vivo. (A) The number of HSCs completing the cell cycle was assessed by in vivo BrdU labeling. HSCs (SP^KLS) were purified from WT and CD48^−/− mice after 3 days of BrdU exposure. (B) HSCs (SP^KLS) from CD48^−/− and WT mice were isolated by flow cytometry and cells were examined for expression of Ki-67, a marker of actively cycling cells. (C) 5-FU was used to challenge the HSCs (SP^SL) in CD48 knockout and WT mice and the cells were examined 6 days post 5-FU. (D) Single HSCs (SP^KLS) were sorted into a 96-well plate with methocult to promote stem cell proliferation and colony formation in 3 separate experiments. Wells were inspected for colonies at regular intervals after plating and the percentage of wells with colonies at each time point was recorded. In this assay, CD48^−/− cells performed equivalently to WT cells.