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. 2005 Feb 1;115(2):268–281. doi: 10.1172/JCI21848

Figure 2.

Figure 2

Characterization of ire1α–/– HSCs. (A) Phenotype of ire1α–/– HSCs from fetal liver and AGM. FACS analysis of CD34 and c-kit in cells from ire1α–/– and ire1α+/+ fetal liver and AGM at E11. The ratio of the number of AGM cells to that of fetal liver cells used for FACS analysis was 2:1. Representative data from 1 of at least 3 separate experiments are shown. (B) BrdU labeling of ire1α–/– and ire1α+/+ fetal livers at E11. The pregnant mouse was injected with 10 mg BrdU at 2 hours before isolation of embryos. Paraffin-embedded sections (5 μM) were stained with anti-BrdU antibody and visualized at ×100 magnification. The proliferating cells in the fetal livers that incorporated BrdU can be identified by dark staining. (C) Proliferation rates of ire1α–/– and ire1α+/– HSCs. Fetal liver cells from WT embryos at E14.5 were first cultured for 2 days to create a hepatic stromal layer. Similar amounts of ire1α–/– and ire1α+/– CD34 and c-kit double-positive HSCs from embryos at E11 were overlaid on the hepatic stromal layer and were cultured for 10 days. The proliferation rates of ire1α–/– and ire1α+/– HSCs were determined by quantification of neomycin gene copies in the genomic DNA isolated from the proliferated ire1α–/– and ire1α+/– cells. The proliferation rate of the ire1α–/– cells was defined as 1. The proliferation rates of other cells were compared with that of the ire1α–/– cells. Experiments were performed at least 3 times, and the SD is indicated.