LPS stimulation induces Bmx kinase activity in primary human macrophages. PBMCs were differentiated into macrophages in the presence of 100 ng/mL of M-CSF for 4 days. Cells were infected for 2 hours with adenoviruses overexpressing HA-tagged wild-type Bmx or control adenovirus Ad0 at a multiplicity of infection of 100:1 in serum-free medium. (A) Expression of HA-Bmx at different multiplicities of infection was assessed by Western blotting. (B) For auto-kinase assay, cells were cultured in complete medium and treated with 10 ng/mL of LPS for 0, 5, 10 and 20 minutes. Cells were lysed and HA-Bmx immunoprecipitated from lysates as described in “Immunoprecipitation and kinase assay” and subjected to in vitro kinase assay. (C) Densitometry units (mean ± SEM) pooled for 3 separate donors are shown normalized to untreated controls. Statistical significance was assessed using one-way ANOVA and Bonferroni multiple comparisons test (**P< .01).