Figure 1

Differentiation of bone marrow cells at different densities results in phenotypically distinct BMDM populations. Plating cells at higher density produces (a) smaller and rounder macrophages, with top panel showing the forward-side scatter plot bottom panel showing the phase contrast images; (b) less CD11b⁺F4/80⁺ cells but more CD11b⁺F4/80⁺CD11c⁺; (c) less CD11b + c-Fms⁺; and (d) more CD11b⁺Ly-6G⁺ cells. For flow cytometric analysis, 2 × 10⁵ BMDM cells detached from culture dishes by 5 mM EDTA 1× PBS were stained with appropriate conjugated primary antibodies, using unstained cells and isotype antibodies as negative controls and Fixable Violet™ staining (Invitrogen) for dead cell exclusion. Data was acquired by BD LSRII flow cytometer and analyzed with Flowjo software. Results represent the mean (±SE) of three experiments of three independent samples each, and statistical analysis was done by student t test with Welch’s correction where appropriate. *** P < 0.0005; ** P < 0.005; * P < 0.05.