Western Blot Analysis of MAPK Signaling Pathway

Western blot was performed as previously described (27 (link)). Samples of equivalent total protein (40 μg) were loaded. Primary antibody against REST, p-Raf-1, p-beta-catenin, p-MEK-2, p-ERK-1/2 (Abcam, Cambridge, UK, 1:500), E-cadherin, N-cadherin, Vimentin, MMP-9, ZO-1, MEK-2, ERK1/2, Raf-1, beta-catenin, beta-Actin, and Histone H3 (ProteinTech Group, Rosemont, USA, 1:500) were used. Suppression of MAPK signaling pathway with U0126 was performed as previously described (28 (link)). In brief, cells incubated in t25 culture flask with 60~70% confluency were treated with 10 μM U0126 for 36 h before further analysis.

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Jin H., Liu P., Kong L., Fei X., Gao Y., Wu T., Sun D, & Tan X. (2019). Identification of RE1-Silencing Transcription Factor as a Promoter of Metastasis in Pancreatic Cancer. Frontiers in Oncology, 9, 291.

Publication 2019

p-Raf-1 p-ERK-1/2 E-cadherin N-cadherin Vimentin ERK1/2 Raf-1 beta-catenin beta-Actin Histone H3

Antibody Beta actin Beta catenin Cells E cadherin Erk1 Histone h3 Mapk signaling pathway Mek 2 Mmp 9 N cadherin Protein Raf 1 U0126 Vimentin Western blot

Corresponding Organization : China Medical University

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Protocol cited in 1 other protocol

Variable analysis

independent variables

Suppression of MAPK signaling pathway with U0126

dependent variables

P-Raf-1
P-beta-catenin
P-MEK-2
P-ERK-1/2
E-cadherin
N-cadherin
Vimentin
MMP-9
MEK-2
ERK1/2
Raf-1
Beta-catenin

control variables

Total protein (40 μg) loaded
Beta-Actin
Histone H3

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