Anti phospho egfr

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Anti-phospho-EGFR is a laboratory reagent used to detect and quantify the phosphorylated form of the Epidermal Growth Factor Receptor (EGFR) protein. It is a specific antibody that binds to the phosphorylated regions of the EGFR protein, allowing for its identification and measurement in various experimental settings.

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Anti-EGFR (81 citations) Phospho-EGFR (7 citations)

3 protocols using anti phospho egfr

Western Blot Protein Analysis Workflow

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Cells were lysed in a RIPA buffer (150 mM NaCl, 50 mM Tris-Cl, 0.1% NaN₃, 1% NP-40, 0.25% sodium deoxycholate, 1 mM EDTA, 1 mM Na₃VO₄, 1 mM NaF and a protease inhibitor mixture). Total protein concentrations were determined using a BCA assay. The total cell lysates were resolved by SDS-PAGE and transferred to a PVDF membrane. The membrane was blocked with 5% non-fat dry milk and probed with antibodies, including anti-phospho-ERK1/2 (Cell Signaling Technology, Beverly, MA, USA), anti-phospho-EGFR (12A3: Santa Cruz Biotechnology, Dallas, TX, USA), anti-TLR2 (Santa Cruz Biotechnology), anti-ERK1/2 (Cell Signaling Technology), and anti-EGFR (1005, Santa Cruz Biotechnology). Signals from western blot analysis were visualized with an ECL detection system (Amersham Pharmacia Biotech, Piscataway, NJ, USA).

Sim M.S., Kim H.J., Jo S.H., Kim C, & Chung I.Y. (2022). Lysophosphatidylserine Induces MUC5AC Production via the Feedforward Regulation of the TACE-EGFR-ERK Pathway in Airway Epithelial Cells in a Receptor-Independent Manner. International Journal of Molecular Sciences, 23(7), 3866.

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Optimizing Epithelial-Mesenchymal Transition Assays

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Catechol (1,2-dihydroxybenzene) was obtained from Sigma Aldrich (St. Louis, MO), prepared 100 mM stock solution in DMSO, and placed in dark at −20°C. Recombinant human epidermal growth factor (EGF) was purchased from Prospec (East Brunswick, NJ), dissolved at a concentration of 100 μg/mL in sterile water containing 0.1% bovine serum albumin (BSA), and placed in deep freezer at −70°C. Polyclonal anti-E-cadherin antibody was purchased from BD Biosciences (San Jose, CA). Polyclonal anti-Vimentin, anti-phospho-EGFR, anti-EGFR, anti-β-actin, anti-CD44, and anti-Nanog antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX). Monoclonal anti-Snail, anti-phospho-AKT, anti-AKT, anti-phospho-ERK, anti-ERK antibodies were obtained from Cell Signaling Technology (Beverly, MA). Horseradish peroxidase-conjugated anti-mouse or anti-rabbit antibodies were obtained from Santa Cruz Biotechnology (Dallas, TX).

Lim W.C., Kim H., Kim Y.J., Jeon B.N., Kang H.B, & Ko H. (2020). Catechol inhibits epidermal growth factor-induced epithelial-to-mesenchymal transition and stem cell-like properties in hepatocellular carcinoma cells. Scientific Reports, 10, 7620.

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Subcellular Fractionation and Western Blot Analysis

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Cells were first lysed in RIPA buffer (50 mM Tris-HCl, 0Á1 M NaCl, 0Á1% NaN 3 , 1% Nonidet P-40, 0Á25% sodium deoxycholate, 1 mM EDTA, 1 mM Na 3 VO 4 , 1 mM NaF and protease inhibitor mixture) for 20 min at 4°a nd then centrifuged for 10 min at 4°. The supernatant represented the cytosolic fraction. The pellet was resuspended with modified RIPA buffer (0Á4 M instead of 0Á1 M NaCl) for 20 min at 4°and centrifuged. The supernatant represented the nuclear fraction. The fractions were resolved by SDS-PAGE and transferred to PVDF membranes. Membranes were blocked with 5% non-fat dry milk and probed with either anti-phospho-ERK1/2 (Cell Signaling Technology, Beverly, MA), anti-phospho-p38 (Santa Cruz Biotechnology, Santa Cruz, CA), antiphospho-JNK (Santa Cruz Biotechnology), anti-phospho-EGFR (12A3: Santa Cruz Biotechnology), anti-p65 NF-jB (Santa Cruz Biotechnology), or anti-IjB (Cell Signaling Technology) antibodies. Anti-ERK2 (C-14: Santa Cruz Biotechnology), anti-p38 (Cell Signaling Technology), anti-JNK (Cell Signaling Technology), anti-EGFR (13G8: Santa Cruz Biotechnology), anti-GAPDH (Santa Cruz Biotechnology), and anti-Histone3 (Abcam) antibodies were used as loading controls. Immunostained proteins were visualized with an ECL detection system (Amersham Pharmacia Biotech, Piscataway, NJ).

Kang J.H., Hwang S.M, & Chung I.Y. (2015). S100A8, S100A9 and S100A12 activate airway epithelial cells to produce MUC5AC via extracellular signal-regulated kinase and nuclear factor-κB pathways. Immunology, 144(1).

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