Rabbit monoclonal anti phospho p38 mapk thr180 tyr182

P1 mice were euthanized by decapitation and bisected heads fixed in 4% PFA in PBS for 1 h at 4°C. The bisected heads were then dehydrated and embedded in paraffin wax and 5-µm sections collected onto charged slides. Sections were de-paraffinised, endogenous peroxidase activity quenched by submersion in 3% H₂O₂, washed in 1× TBST and blocked in 1× TBST containing normal goat serum. Sections were then incubated overnight at 4°C with rabbit monoclonal anti-phospho p38 MAPK (Thr180/Tyr182) (Cell Signaling Technology) at 1:1500 dilution, phospho-p44/42 MAPK (ERK1/2) (Cell Signaling Technology) at 1:1000 dilution or phospho-SAPK/JNK (Thr183/Tyr185) (Cell Signaling Technology) at 1:1000 dilution. The VECTASTAIN^® Elite ABC rabbit IgG avidin biotin kit (Vector Laboratories) and DAB+ Chromagen (Dako) were used for detection. For anti-phospho-p38-abelled images, the ImmunoRatio plugin (http://jvsmicroscope.uta.fi/immunoratio/) for ImageJ (http://imagej.nih.gov/ij/) was used to quantify the percentage of positively stained nuclei.

Western blot analysis was performed according to the description previously [20 (link)]. The following primary antibodies were used: rabbit polyclonal anti-ABCB1 (1: 500, Proteintech, Rosemont, USA), rabbit monoclonal anti-CCR7 (1:10,000, Abcam, Cambridge, MA), mouse monoclonal anti-ERK1/2 (1:2000, Proteintech), mouse monoclonal anti-P38 MAPK (1: 2000, Proteintech). rabbit monoclonal anti-phospho-p44/42 MAPK(Erk1/2)(Thr202/Tyr204, 1: 1000, Cell Signaling Technology, Danvers, MA, USA), rabbit monoclonal anti-phospho-p38 MAPK (Thr180/Tyr182, 1: 1000, Cell Signaling Technology), mouse monoclonal anti-E-cadherin (1:2000, Proteintech), mouse monoclonal anti-Vimentin(1:5000, Proteintech), rabbit polyclonal anti-VEGF(1: 1000, Proteintech). Rabbit polyclonal anti-GAPDH (1:5000, Proteintech) was used as an internal control.