Egfr sc 03

Manufactured by Santa Cruz Biotechnology

Sourced in United States

EGFR (sc-03) is a laboratory reagent produced by Santa Cruz Biotechnology. It is an antibody that can be used to detect the presence of the epidermal growth factor receptor (EGFR) protein in biological samples.

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Close spelling variants of this product

Sc-03 (21 citations) Anti-EGFR (sc-03) (15 citations) Anti-EGFR antibody (SC-03) (5 citations) Anti-EGFR ((1005) SC-03 – (4 citations) Total EGFR antibody (Sc-03) (3 citations) EGFR antibody (sc-03) (2 citations) EGFR (sc-03) and ERK1 (sc-94) (2 citations) EGFR (sc‐03‐G, (2 citations)

8 protocols using egfr sc 03

Visualizing EGFR and EEA1 in Transfected Cells

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Cells transfected with scrambled siRNA or siRNA targeting GSDME were seeded on coverslips for the indicated time, washed with PBS three times and fixed in 4% paraformaldehyde for 15 min. Subsequently, the cells were permeabilized in 0.2% Triton X-100 for 10 min, blocked in 10% normal goat serum for 1 h at 4 °C and incubated with EEA1 (E7659, Sigma‒Aldrich) and EGFR (sc-03, Santa Cruz) antibodies overnight at 4 °C. The next day, following washing with PBS three times, Alexa Fluor 488 and Alexa Fluor 555 secondary antibodies were added to the sections and incubated for 1 h at room temperature. The sections were washed in PBS three times and counterstained with DAPI. Fluorescence images were visualized with a laser scanning confocal microscope.

Xu L., Shi F., Wu Y., Yao S., Wang Y., Jiang X., Su L, & Liu X. (2023). Gasdermin E regulates the stability and activation of EGFR in human non-small cell lung cancer cells. Cell Communication and Signaling : CCS, 21, 83.

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Protein Isolation and Immunoblotting Assay

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Protein was isolated in RIPA buffer (Millipore), supplemented with Halt protease inhibitor (Thermo Scientific, Rockford, IL, USA). Primary antibodies were used according to the manufacturer's instructions: TFAP2C #sc-12762, NEU #sc-284, ERK #sc-93, GAPDH #sc-32233 and EGFR sc-03 (Santa Cruz Biotechnology, Santa Cruz, CA, USA); pERK #4370 (Cell Signaling, Danvers, MA, USA). Secondary antibodies were used according to manufacturer's instructions: anti-rabbit HRP #sc-2030 and anti-mouse HRP #sc-2005. Protein was visualized with SuperSignal West Dura extended duration substrate (Thermo Scientific) and SuperSignal West Femto maximum sensitivity substrate (Thermo Scientific).

Park J., Wu T., Cyr A., Woodfield G., De Andrade J., Spanheimer P., Li T., Sugg S., Lal G., Domann F., Zhang W, & Weigel R. (2015). The role of Tcfap2c in tumorigenesis and cancer growth in an activated Neu model of mammary carcinogenesis. Oncogene, 34(50), 6105-6114.

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Estrogen Receptor Signaling Pathway

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17β-estradiol (E2) was purchased from Sigma-Aldrich (St. Louis, MO). Recombinant hPRL (Lot AFP795) was obtained from Dr. A.F. Parlow (National Hormone and Pituitary Program, NIDDK, National Institutes of Health, Torrance, CA). Type-I rat tail collagen (#CB354249) was obtained from Fisher Scientific (Pittsburgh, PA). Inhibitors used for these studies were purchased as follows: 4-hydroxy-tamoxifen (4-OHT) (#579002) from EMD Millipore (Billerica, MA), SFK inhibitor, PP-1 (#EI275) from Biomol International, LP (Plymouth Meeting, PA), and JAK2 inhibitor, BMS-911543 (#CT-BMS91) from Chemietek (Indianapolis, IN). Type-I collagenase (#17100–017) was purchased from Invitrogen (Grand Island, NY). Antibodies used in these studies were as follows: PRLR-ECD (#35–9200) and pSRC Y418 (#44660G) from Invitrogen (Grand Island, NY); ERK1/2 (#9102) and cleaved caspase-3 (#9661) from Cell Signaling Technology (Danvers, MA); cSRC (sc-18) and EGFR (sc-03) from Santa Cruz Biotechnology (Santa Cruz, CA); ERα (#NCL-ER-6F211/2) from Novocastra (Newcastle, United Kingdom); pan-actin (#125-ACT) from Phosphosolutions (Aurora, CO); Ki-67 (#AB15580) from Abcam (Cambridge, MA). Multiwell non-tissue culture-treated plates were obtained from Corning Life Sciences (#08–772–49 and #08–772–51, Tewksbury, MA). All other reagents were obtained from Fisher Scientific or Sigma-Aldrich.

Barcus C.E., Holt E.C., Keely P.J., Eliceiri K.W, & Schuler L.A. (2015). Dense Collagen-I Matrices Enhance Pro-Tumorigenic Estrogen-Prolactin Crosstalk in MCF-7 and T47D Breast Cancer Cells. PLoS ONE, 10(1), e0116891.

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Automated Capillary-Based Protein Quantification

Cited 2 times

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Cell pellets were lysed in 50 mM Tris buffer (pH 8.0), 150 mM NaCl, 1% Triton X-100, protease inhibitors (Complete Mini, Roche) and anti-phosphatases (PhosSTOP, Roche) for 1 h at 4 °C. Lysates were centrifuged for 20 min at 15000 × g at 4 °C. Protein expression studies were performed by WES, an automated capillary-based size sorting system (ProteinSimple, San Jose, CA, USA)^{33 (link),34 (link)}. Data were analyzed using Compass software (ProteinSimple, San Jose, CA, USA). The primary antibodies used were EGFR (sc-03; Santa Cruz Biotechnology, Santa Cruz, CA, USA), phospho-EGFR (Tyr1068; Cell Signaling Technology, Danvers, MA, USA), HER2 (Cell Signaling Technology), HER3 (Cell Signaling Technology), phospho-HER2 (Y1221/1222) (Cell Signaling Technology), phospho-HER3 (Y1283) (Cell Signaling Technology), phospho-AKT (Ser473; Cell Signaling Technology) and phospho-MEK1/2 (Ser217/221; Cell Signaling Technology) at a 1/50 dilution, and GAPDH (Santa Cruz Biotechnology) was used as a reference. Protein expressions are represented as digitized images of blotting by quantitative chemiluminescence. Each experiment was performed in triplicate.

Guy J.B., Méry B., Ollier E., Espenel S., Vallard A., Wozny A.S., Simonet S., Lauret A., Battiston-Montagne P., Ardail D., Alphonse G., Rancoule C., Rodriguez-Lafrasse C, & Magné N. (2017). Dual “mAb” HER family blockade in head and neck cancer human cell lines combined with photon therapy. Scientific Reports, 7, 12207.

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Western Blot Analysis of Signaling Pathways

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Cells were harvested and washed in PBS and lysed in RIPA buffer (9806, Cell Signaling Technology). Western blot analysis was performed by using the conventional protocols as described previously (Ma et al., 2016 (link)). Primary antibodies were used including EGFR (sc-03, Santa Cruz Biotechnology), phospho-EGFR (Tyr1068) (3777, Cell Signaling Technology), phospho-Stat3 (Tyr705) (9145, Cell Signaling Technology), Stat3 (9132, Cell Signaling Technology), phospho-Akt (Ser473) (9271, Cell Signaling Technology), Akt (9272, Cell Signaling Technology), GAPDH (60004-1-Ig, Proteintech). After extensively washed, the membranes were then incubated with secondary antibodies (Zhongshan Golden Bridge Biotechnology Company, Beijing, China) for 1 h at room temperature. Signals were detected using enhanced chemiluminescence (Engreen, Beijing, China). The bands intensities were analyzed using ImageJ software.

Wang Y., Liu X., Hu G., Hu C., Gao Y., Huo M., Zhu H., Liu M, & Xu N. (2021). EGFR-IL-6 Signaling Axis Mediated the Inhibitory Effect of Methylseleninic Acid on Esophageal Squamous Cell Carcinoma. Frontiers in Pharmacology, 12, 719785.

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Immunohistochemistry of Tumor Markers

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IHC was conducted as described previously [48 (link)]. Following primary antibodies were used: anti-pan cytokeratin (1:200, pan-CK, NB120-6401, Novus biological, Littleton, CL, USA), anti-epidermal growth factor receptor (1:30, EGFR, sc-03, Santa Cruz, Santa Cruz, CA, USA), and anti-Ki-67 (1:500, NCL-Ki-67p, Novocastra laboratories, Newcastle upon Tyne, UK).

Chen Y.L., Liu K.J., Jang C.W., Hsu C.C., Yen Y.C., Liu Y.L., Chuang T.H., Wang S.H., Fu Y.K., Kuo C.C, & Chen Y.W. (2019). ERK Activation Modulates Cancer Stemness and Motility of a Novel Mouse Oral Squamous Cell Carcinoma Cell Line. Cancers, 12(1), 61.

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Antibody Sources for Signaling Pathway Analysis

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Antibodies were obtained from the following sources: Antibodies against phospho-p65 (CST-3033), p65 (CST-6956), phospho-IKKα/β (CST-2697), IKKα (CST-2682), IKKβ (CST-8943), phospho-Akt (CST-4508), Akt1 (CST-2938), Akt2 (CST-3063), phospho-S6K (CST-9205), mTOR (CST-2972), cleaved caspase 3 (CST-9664), and GAPDH (CST-5174), are purchased from Cell Signaling. Anti-Raptor ((A300-506A) and anti-Rictor (A300-458A) are from Bethyl. Anti-HA (H6908) is from Sigma. Anti-S6K (SC-8418), C-Myc (SC-40), and EGFR (SC-03) are from Santa Cruz Biotechnology. HRP-labeled anti-mouse and anti-rabbit secondary antibodies were also from Santa Cruz Biotechnology.

Li Z., Yang Z., Passaniti A., Lapidus R.G., Liu X., Cullen K.J, & Dan H.C. (2016). A positive feedback loop involving EGFR/Akt/mTORC1 and IKK/NF-κB regulates head and neck squamous cell carcinoma proliferation. Oncotarget, 7(22), 31892-31906.

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Western Blot Analysis of NRG1 Signaling

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Cells were washed twice in cold PBS and lysed with Laemmeli sample buffer. Lysates were resolved by SDS polyacrylamide gel electrophoresis and transferred to PVDF membranes. For secreted NRG1 detection, medium was collected and centrifuged for 30 mins at 4000 rpm in an Amicon ultra conical tube. Laemmeli sample buffer was added and samples resolved by SDS-PAGE. After blocking in 5% BSA, membranes were incubated with the indicated primary antibodies overnight at 4 °C, followed by incubation with peroxidase-coupled secondary antibodies. Bound antibodies were detected using enhanced chemiluminescence substrate (Pierce, Rockford IL). Chemiluminescence was visualized on a VersaDoc Multi-Imager and quantitated using Quantity-One software (BioRad, Hercules, CA). Primary antibodies used were: NRG1 (# MAB377) antibody purchased from R&D Biosystems (Minneapolis, MN) used for detection of secreted NRG1, ERK2 (sc-1647) and EGFR (sc-03) antibodies purchased from Santa Cruz Biotechnology Inc. (Santa Cruz, CA); NRG1 (#2573), p-ErbB3 (Y1197, #4561), ErbB3 (#4754), p-ErbB2 (Y1196, #6942), ErbB2 (#4290), p-AKT (S473, #6942), p-AKT (T308, #2965), AKT (#9272) and p-ERK1/2 (T202/Y204, #9101) purchased from Cell Signaling Technology; actin (A2066) antibody purchased from Sigma-Aldrich Co. (St. Louis, MO).

Capparelli C., Rosenbaum S., Berman-Booty L.D., Salhi A., Gaborit N., Zhan T., Chervoneva I., Roszik J., Woodman S.E., Davies M.A., Setiady Y.Y., Osman I., Yarden Y, & Aplin A.E. (2015). ErbB3/ErbB2 complexes as a therapeutic target in a subset of wild-type BRAF/NRAS cutaneous melanomas. Cancer research, 75(17), 3554-3567.

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