Jak2^FF/FFleads to partial IFN-γ signaling in primary macrophages and MEFs. (A) Jak2^+/+, Jak2^FF/FF, and Jak2^−/− embryos (upper panel) and embryonic livers (lower panel) derived at E12.5. (B) Embryonic liver cells from Jak2^+/+, Jak2^+/FF, and Jak2^FF/FF embryos were prepared at E12.5 and stimulated for 1 hour with Epo or left untreated. Cellular lysates were prepared and analyzed by immunoblotting using the indicated antibodies. (C) Fetal liver cells from Jak2^+/+, Jak2^FF/FF, and Jak2^−/− embryos were prepared at E12.5 and were incubated with macrophage colony-stimulating factor for 6 days. FLDM were then stimulated with IFN-γ for 48 hours or left untreated. Cells were stained for F4/80 as a marker for mature macrophages. F4/80-positive cells were analyzed for MHC-I (H-2K^b) surface expression as a marker for IFN-γ signaling. One representative of 3 independent experiments run in triplicate is shown. The MHC-I expression of each genotype without stimulation was set as 100%. The standard deviation was below 3.7% of the detected values. ***P < .001. (D) MEFs from Jak2^+/+, Jak2^FF/FF, and Jak2^−/− mice were stimulated in vitro with IFN-γ for the indicated time points. Subsequently, cellular lysates were prepared and analyzed by immunoblotting for expression of phospho-Stat1 (Y701), Stat1, and Jak2. β-actin served as a loading control. Results shown are representative of 3 independent experiments.