Isolation and characterization of different stromal cell populations from mouse fetal livers. Fetal liver cell suspensions obtained following enzymatic digestion of dissected E13.5 fetal liver were subjected to fluorescence-activated cell sorting analysis and CFU-F assays. (A) Fluorescence-activated cell sorting analysis before (i) and after (ii) magnetic depletion of CD45/TER119 hematopoietic cells, and the gating strategy used for the sorting of the different cell populations tested for CFU-F content (iii-iv). Dot plots and histograms are representative figures with the mean ± standard error of the mean (SEM) from 27 independent experiments. (B) Sorted cell populations were plated in α-minimal essential medium 20% fetal calf serum, and CFU-F frequencies were determined in limiting dilution assays using L-Calc software. Values represent mean ± SEM from 3 independent experiments. (C) Hem⁻CD51⁺VCAM-1⁺PDGFRα⁻ (V⁺P⁻) cells that failed to produce CFU-F were sorted and plated in EGM-2. (i) Phase contrast images of the adherent layers constituted of large spread cells. Scale bar represents 40 μm. (ii) Expression of E-cadherin (E-Cadh), c-met, epithelial cell adhesion molecule (EpCAM), and Dlk-1 for the V⁺P⁻ cell population. Shown is a representative histogram from 3 independent experiments. All histogram markers are based upon fluorescence-minus-one controls. (iii) Expression of Hnf4a, Alb, Afp, Krt18, c-met, G6pc, and GAPDH transcripts in sorted cells from the fetal liver and total adult liver. Figures on the left side of the panel indicate amplicon size. FL, fetal liver; MW, molecular weight; Neg, negative.