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Anti phospho egf receptor tyr1068

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Anti-phospho-EGF-Receptor Tyr1068 is a laboratory reagent that detects the phosphorylation of the epidermal growth factor (EGF) receptor at tyrosine residue 1068. It is used as a research tool to study the activation and signaling of the EGF receptor in biological samples.

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Lab products found in correlation

Anti phospho met tyr1234 y1235, by Cell Signaling Technology (4 mentions) Anti her3 d22c5 xp, by Cell Signaling Technology (4 mentions) Widefield epifluorescent microscope, by Nikon (2 mentions) Alexa fluor 488 donkey anti rabbit, by Thermo Fisher Scientific (2 mentions) Anti phospho her3 tyr1289, by Cell Signaling Technology (2 mentions) Anti met d1c2, by Cell Signaling Technology (2 mentions) Anti golgin 97, by Thermo Fisher Scientific (2 mentions) Anti pdi rl90, by Thermo Fisher Scientific (2 mentions) Alexa fluor 568 goat anti mouse, by Thermo Fisher Scientific (2 mentions) Anti flag m2 antibody, by Merck Group (2 mentions) Anti eea1, by BD (2 mentions) Spinning disc confocal microscope, by Nikon (2 mentions) β tubulin 9f3, by Cell Signaling Technology (2 mentions) Anti met d1c2 xp, by Cell Signaling Technology (2 mentions) Anti her3 sc 285, by Santa Cruz Biotechnology (2 mentions) Anti her3 sc 81455, by Santa Cruz Biotechnology (2 mentions) Anti phospho her3 tyr1289 21d3, by Cell Signaling Technology (2 mentions) Anti her2 sc 284, by Santa Cruz Biotechnology (2 mentions) Anti phospho her2 tyr1221 1222, by Cell Signaling Technology (2 mentions) Anti egfr 1005 sc 03, by Santa Cruz Biotechnology (2 mentions)

Spelling variants of this product

EGF Receptor (9 citations) Phospho-EGF Receptor (Tyr1068) (6 citations) Phospho-EGF Receptor (5 citations) Anti-EGF Receptor (4 citations) Anti-phospho-EGF receptor (2 citations) Rabbit anti-Phospho-EGF Receptor (Tyr1068) (2 citations)

5 protocols using anti phospho egf receptor tyr1068

1

Quantifying Receptor Localization and Activation

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9×104 COS7 cells were plated on coverslips. After 24 hours, cells were transfected and cultured for 24–48 hours. Cells were fixed with 3.7% formaldehyde for 1 hour at room temperature, permeabilized with 0.1% Triton X-100 for 5 minutes, and blocked with 1% BSA for 5 minutes. Cells were incubated with primary antibodies at 37C for 1–2 hours: anti-FLAG-M2 antibody (Sigma Aldrich #F1804), anti-phospho-HER3 Tyr1289 (21D3 – Cell Signaling), anti-MET (D1C2 – Cell Signaling), anti-phospho-MET Tyr1234/Y1235 (Cell Signaling), anti-phospho-EGF-Receptor Tyr1068 (Cell Signaling), anti-Golgin97 (A-21270 – Molecular Probes), anti-PDI (RL90 - Thermo Scientific), or anti-EEA1 (BD Biosciences - 610456), anti-HER3 (D22C5 XP – Cell Signaling). Cells were washed and incubated with fluorescently tagged anti-rabbit or anti-mouse secondary antibodies (Goat Anti-Mouse Alexa-fluor-568 cat#A11031; Alexa-Fluor-488 donkey anti-rabbit cat# A21206 – Life technologies). Images were taken at 60x magnification using a Nikon widefield epifluorescent microscope (Fig. 3a, b, 5a-d, 7b), or at 60x using a Nikon spinning disc confocal microscope Confocal images shown were taken as a stack of 7 slices in the Z-plane and processed as a Z-max projection (Supplementary Fig. 4).
Frazier N.M., Brand T., Gordan J.D., Grandis J, & Jura N. (2018). Overexpression-mediated activation of MET in the Golgi promotes HER3/ERBB3 phosphorylation. Oncogene, 38(11), 1936-1950.
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2

Western Blot Analysis of Receptor Tyrosine Kinases

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Cells were lysed in lysis buffer (50mM Tris pH 7.5, 150mM NaCl, 1mM EDTA, 1mM Na3VO4, 1mM NaF, 1% Triton X-100), or RIPA buffer (50mM Tris pH 7.5, 150mM NaCl, 1mM EDTA, 1mM Na3VO4, 1mM NaF, 0.1% SDS, 0.1% deochycholate, 1% IGEPAL CA-630) with protease inhibitor cocktail (Roche). Lysates were run on 8 or 10% SDS-PAGE and transferred to PVDF membrane (EMD Millipore). Proteins were detected using: anti-MET (D1C2 XP– Cell Signaling), anti-phospho-MET Tyr1234/Y1235 (Cell Signaling), anti-HER3 (SC-285 – Santa Cruz), anti-HER3 (SC-81455 – Santa Cruz), anti-HER3 (D22C5 XP – Cell Signaling), anti-phospho-HER3 Tyr1289 (21D3 – Cell Signaling), anti-HER2 (SC-284 – Santa Cruz), anti-phospho-HER2 Tyr1221/1222 (Cell Signaling), anti-FLAG M2 (Sigma-Aldrich), anti-EGFR ((1005) SC-03 – Santa Cruz), anti-phospho-EGF-Receptor Tyr1068 (Cell Signaling), β-tubulin (9F3 – Cell Signaling), GAPDH (D4C6R - Cell Signaling). Secondary antibodies were anti-rabbit-IgG HRP-linked antibody (Cell Signaling), or anti-mouse IgG HRP-linked whole antibody (GE Healthcare Biosciences). Blots were developed using ECL/ECL Prime (Thermo Fisher Scientific).
Frazier N.M., Brand T., Gordan J.D., Grandis J, & Jura N. (2018). Overexpression-mediated activation of MET in the Golgi promotes HER3/ERBB3 phosphorylation. Oncogene, 38(11), 1936-1950.
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3

Western Blot Analysis of Receptor Tyrosine Kinases

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Cells were lysed in lysis buffer (50mM Tris pH 7.5, 150mM NaCl, 1mM EDTA, 1mM Na3VO4, 1mM NaF, 1% Triton X-100), or RIPA buffer (50mM Tris pH 7.5, 150mM NaCl, 1mM EDTA, 1mM Na3VO4, 1mM NaF, 0.1% SDS, 0.1% deochycholate, 1% IGEPAL CA-630) with protease inhibitor cocktail (Roche). Lysates were run on 8 or 10% SDS-PAGE and transferred to PVDF membrane (EMD Millipore). Proteins were detected using: anti-MET (D1C2 XP– Cell Signaling), anti-phospho-MET Tyr1234/Y1235 (Cell Signaling), anti-HER3 (SC-285 – Santa Cruz), anti-HER3 (SC-81455 – Santa Cruz), anti-HER3 (D22C5 XP – Cell Signaling), anti-phospho-HER3 Tyr1289 (21D3 – Cell Signaling), anti-HER2 (SC-284 – Santa Cruz), anti-phospho-HER2 Tyr1221/1222 (Cell Signaling), anti-FLAG M2 (Sigma-Aldrich), anti-EGFR ((1005) SC-03 – Santa Cruz), anti-phospho-EGF-Receptor Tyr1068 (Cell Signaling), β-tubulin (9F3 – Cell Signaling), GAPDH (D4C6R - Cell Signaling). Secondary antibodies were anti-rabbit-IgG HRP-linked antibody (Cell Signaling), or anti-mouse IgG HRP-linked whole antibody (GE Healthcare Biosciences). Blots were developed using ECL/ECL Prime (Thermo Fisher Scientific).
Frazier N.M., Brand T., Gordan J.D., Grandis J, & Jura N. (2018). Overexpression-mediated activation of MET in the Golgi promotes HER3/ERBB3 phosphorylation. Oncogene, 38(11), 1936-1950.
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4

Quantifying Receptor Localization and Activation

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9×104 COS7 cells were plated on coverslips. After 24 hours, cells were transfected and cultured for 24–48 hours. Cells were fixed with 3.7% formaldehyde for 1 hour at room temperature, permeabilized with 0.1% Triton X-100 for 5 minutes, and blocked with 1% BSA for 5 minutes. Cells were incubated with primary antibodies at 37C for 1–2 hours: anti-FLAG-M2 antibody (Sigma Aldrich #F1804), anti-phospho-HER3 Tyr1289 (21D3 – Cell Signaling), anti-MET (D1C2 – Cell Signaling), anti-phospho-MET Tyr1234/Y1235 (Cell Signaling), anti-phospho-EGF-Receptor Tyr1068 (Cell Signaling), anti-Golgin97 (A-21270 – Molecular Probes), anti-PDI (RL90 - Thermo Scientific), or anti-EEA1 (BD Biosciences - 610456), anti-HER3 (D22C5 XP – Cell Signaling). Cells were washed and incubated with fluorescently tagged anti-rabbit or anti-mouse secondary antibodies (Goat Anti-Mouse Alexa-fluor-568 cat#A11031; Alexa-Fluor-488 donkey anti-rabbit cat# A21206 – Life technologies). Images were taken at 60x magnification using a Nikon widefield epifluorescent microscope (Fig. 3a, b, 5a-d, 7b), or at 60x using a Nikon spinning disc confocal microscope Confocal images shown were taken as a stack of 7 slices in the Z-plane and processed as a Z-max projection (Supplementary Fig. 4).
Frazier N.M., Brand T., Gordan J.D., Grandis J, & Jura N. (2018). Overexpression-mediated activation of MET in the Golgi promotes HER3/ERBB3 phosphorylation. Oncogene, 38(11), 1936-1950.
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5

Immunofluorescence Analysis of Stem Cell Markers

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Cells plated on coverslips were fixed in 4% paraformaldehyde (w/v) for 15 min, and permeabilized in PBS/0.3% Triton X-100 for another 15 min. Cells were then washed with PBS and blocked in 2% BSA for 1 h. Primary antibodies namely, anti-CD109 (C-9) (Santa Cruz Biotechnology, Santa Cruz, CA, USA; sc-271085), anti-EGFR (Cell Signaling, Danvers, MA, USA; mAb #2232), anti-Phospho-EGF Receptor (Tyr1068) (Cell Signaling, Danvers, MA, USA; mAb #3777), anti-Sox2 (cell Signaling Danvers, MA, USA; mAb #3108), anti-Oct4 (Santa Cruz, Santa Cruz, CA, USA; sc-23900), and anti-NANOG (Santa Cruz, Santa Cruz, CA, USA; sc-23900) antibodies were then added to cells at 1:300 dilution in 2% BSA and incubated overnight at 4°C. Cells were washed with PBS and labeled for 1 h with fluorophore-conjugated secondary antibodies (1:500 dilution) specifically, Alexa Fluor 594-goat anti-rabbit (Life Technology; A11037) and Alexa Fluor 488 goat-anti-mouse (Life Technology; A11029). Cells were washed with PBS, and mounted with Fluoroshield mounting medium with DAPI (Abcam; ab104139). Cells were visualized using an Olympus microscope IX71.
Zhou S., Hassan A., Kungyal T., Tabariès S., Luna J.L., Siegel P.M, & Philip A. (2022). CD109 Is a Critical Determinant of EGFR Expression and Signaling, and Tumorigenicity in Squamous Cell Carcinoma Cells. Cancers, 14(15), 3672.
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