Analysis of myeloid lineage properties and in vitro differentiation potential. (A) Sorted B220⁺Mac1⁻/Gr-1⁻ cells were evaluated for the ability to generate myeloid colonies in M3434 methylcellulose. (B) MPO expression in the B220⁺Mac-1⁻/Gr-1⁻, compared with WT controls. (C) MPO expression in the B220⁺Mac-1⁺/Gr-1⁺ population compared with WT controls. (D) B220⁺Mac-1⁻/Gr-1⁻ cells from FLT3/ITD-NHD13 mice were grown in liquid culture with myeloid cytokines and evaluated for differentiation ability and (E) functional activity. Images were acquired at room temperature using a Nikon TE 2000-E microscope system (Nikon) with a Nikon Plan APO VC 100×/1.40 oil objective and Nikon EZ-C1 Version 3.5 software. (F) Cells differentiated in liquid culture were stained for markers of macrophage differentiation.