ERK1/2 Antibody

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	-Rabbit (1:3000) -Soak PVDF membranewith pore size 0.45 um in methanol for 30 sec -Apply Wet transfersystems at 180mA for 90 min with Wet transfer buffer.

Protocol tips
-Rabbit (1:3000) -Soak PVDF membranewith pore size 0.45 um in methanol for 30 sec -Apply Wet transfersystems at 180mA for 90 min with Wet transfer buffer.

Publication protocol

Cells or tissues from mice were lysed in SDS lysis buffer (2% SDS and 62.5 mM Tris-HCl pH 6.8, containing protease inhibitor cocktail tablets ordered from Roche) and sonicated three times for 5 s. Equal amounts of protein (20 μg) were loaded onto a 10% polyacrylamide gel and analyzed by immunoblotting. The antibodies used for western blotting were against VEGFR2 [Cat# 2479S, Cell Signaling Technology (CST), Danvers, MA,, 1:2000 dilution], phosphorylated (p)-VEGFR2 (Cat# 2478S, CST, 1:2000 dilution), ERK1/2 (Cat# 16443-1-AP, Proteintech, Wuhan, China, 1:3000 dilution), p-ERK1/2 (Cat# 4370S, CST, 1:2000 dilution), MEK1 (Cat# 2352S, CST, 1:2000 dilution), p-MEK1/2 (Cat# 2338S, CST, 1:2000 dilution), p-PLCγ (Cat# 8713T, CST, 1:2000 dilution), SRC (Cat# 2123T, CST, 1:2000 dilution), p-SRC (Cat# 6943T, CST, 1:2000 dilution), p38 MAPK family proteins (Cat# 8690T, CST, 1:2000 dilution), p-p38 MAPK family proteins (Cat# 4511T, CST, 1:2000 dilution), AKT (Cat# 10176-2-AP, Proteintech, 1:2000 dilution), p-AKT (Cat# 4060S, CST, 1:2000 dilution) and GAPDH (Cat# 60004-1-Ig, Proteintech, 1:3000 dilution). HRP-conjugated goat anti-rabbit-IgG secondary antibody (Cat# 7074, CST, 1:5000 dilution) and goat anti-mouse-IgG secondary antibody (Cat# 7076, CST, 1:5000 dilution) were used for immunoblotting.



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