Fig. 2
From: Bruton’s tyrosine kinase is a possible therapeutic target in microscopic polyangiitis
Btk and Vav phosphorylation in MPO and anti-MPO-IC-stimulated neutrophils. Human peripheral blood neutrophils were applied to two-well chambers on MPO and anti-MPO-IC-immobilized slides (1 × 106/well) and allowed to settle at 37 °C. As controls, neutrophils were applied to two-well chambers on IC-nonimmobilized or anti-MPO antibody-immobilized slides (1 × 106/well). Three minutes later, neutrophil lysates were collected and subjected to western blotting using anti-Btk and anti-phosphorylated Btk (p-Btk) antibodies (a) and anti-Vav and anti-phosphorylated Vav (p-Vav) antibodies (b). Because anti-Btk and anti-Vav antibodies react with both nonphosphorylated and phosphorylated antigens, each specific band was labeled as total Btk (t-Btk) and total Vav (t-Vav). Experiments were repeated thrice, the intensity of specific bands (arrowheads) was quantified using ImageJ, and the phosphorylation degree in the samples was calculated as p-Btk/t-Btk and p-Vav/t-Vav values. The value of unstimulated neutrophils was set as 1, and the relative values of MPO and anti-MPO-IC-stimulated neutrophils were shown in the graphs with error bars indicating standard deviations. *p < 0.05