Western Blot Analysis of Signaling Proteins

Total cell lysates were collected as preciously described [38 (link)]. The lysates were then incubated overnight at 4 °C with the primary antibodies and then with the secondary antibodies for 1 h at room temperature. Antibodies used were as follows: anti-CAIX (#5649, Cell Signaling), anti-p-ERK (#4370, Cell Signaling), anti-ERK (#9102, Cell Signaling), anti-E-cadherin (610182, BD), anti-vimentin (sc-6260, Santa Cruz), anti-PFKFB4 (GTX 107755, GeneTex), anti-p-MEK (#9121, Cell Signaling), anti-p-C-Raf (#9427, Cell Signaling), anti-C-Raf (#9422, Cell Signaling), and anti-actin (ab8226, Abcam).

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Hsin M.C., Hsieh Y.H., Hsiao Y.H., Chen P.N., Wang P.H, & Yang S.F. (2021). Carbonic Anhydrase IX Promotes Human Cervical Cancer Cell Motility by Regulating PFKFB4 Expression. Cancers, 13(5), 1174.

Publication 2021

anti-p-ERK anti-ERK anti-E-cadherin anti-vimentin anti-p-MEK anti-p-C-Raf anti-C-Raf anti-actin

Actin Antibodies Cadherin Caix Cell Vimentin

Corresponding Organization : Chung Shan Medical University Hospital

Other organizations : Changhua Christian Hospital

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Variable analysis

independent variables

Incubation of total cell lysates with primary antibodies overnight at 4 °C
Incubation of total cell lysates with secondary antibodies for 1 hour at room temperature

dependent variables

Expression levels of CAIX, p-ERK, ERK, E-cadherin, vimentin, PFKFB4, p-MEK, p-C-Raf, and C-Raf

control variables

Total cell lysates were collected as previously described [38]

controls

No positive or negative controls were explicitly mentioned.

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