Western Blot Analysis of Macrophage Proteins

Whole cell proteins were extracted from macrophages using a lysis buffer (Cell Signaling, Cat. No. 9803). Each sample was applied into 10% SDS-PAGE and transferred to a polyvinylidene fluoride membrane, immunoblotted with primary antibodies (the phosphorylation of extracellular signal-regulated kinase (p-ERK); Cell Signaling, Cat. No. 9101, the phosphorylation of c-Jun N-terminal kinase (p-JNK); Cell Signaling, Cat. No. 9251, the urokinase plasminogen activator (uPA); Abcam, Cat. No. ab20789, the urokinase plasminogen activator and receptor (uPAR); R&D, Cat. No. AF534, and β-actin; Sigma Cat. No. A5441) [28 (link)]. Membranes were then incubated with appropriate secondary antibodies, and immune complexes were visualized on chemiluminescence (Merck Millipore, Cat. no. WBLUF0100, Cat. no. WBLUC0100) and quantified using a General Electric Imager (GE Healthcare, LAS 4000 mini) [29 (link)].