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      Alexa Fluor® 647 Mouse anti-EGF Receptor (pY845)
      Alexa Fluor® 647 Mouse anti-EGF Receptor (pY845)
      Analysis of EGF Receptor (pY845) in human epidermoid carcinoma.  A-431 cells were either treated for 30 minutes with EGF (Cat. No. 354052 or 356052, shaded histogram) or untreated (open histogram).  The cells were fixed (BD Cytofix™ buffer, Cat. No. 554655) for 10 minutes at 37°C, then permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, and then stained with Alexa Fluor® 647 Mouse anti-EGF Receptor (pY845).  Flow cytometry was performed on a BD FACSArray™ bioanalyzer system.
      Alexa Fluor® 647 Mouse anti-EGF Receptor (pY845)
      Analysis of EGF Receptor (pY845) in human epidermoid carcinoma.  A-431 cells were either treated for 30 minutes with EGF (Cat. No. 354052 or 356052, shaded histogram) or untreated (open histogram).  The cells were fixed (BD Cytofix™ buffer, Cat. No. 554655) for 10 minutes at 37°C, then permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, and then stained with Alexa Fluor® 647 Mouse anti-EGF Receptor (pY845).  Flow cytometry was performed on a BD FACSArray™ bioanalyzer system.
      Product Details
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      BD Phosflow™
      Human (QC Testing), Mouse,Rat (Reported)
      Mouse IgG1, κ
      Phosphorylated Human EGF Receptor Peptide
      Intracellular staining (flow cytometry) (Routinely Tested)
      20 µl
      AB_647124
      Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
      RUO


      Preparation And Storage

      Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.
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      Product Notices

      1. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
      2. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
      3. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
      4. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
      5. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
      6. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
      7. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      8. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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      558523 Rev. 4
      Antibody Details
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      12A3

      Epidermal Growth Factor (EGF) elicits a variety of cellular responses that are initiated by EGF Receptor (EGFR) binding and activation of intrinsic tyrosine kinase activity.  EGFR, also known as ErbB1 or HER1, is a member of the ErbB class of receptor protein tyrosine kinases.  It has an extracellular ligand-binding domain, a single transmembrane region, and a cytoplasmic region containing a protein tyrosine kinase domain and a c-terminal regulatory domain with many phosphorylation sites.  Following ligand binding, EGFR forms homodimers and heterodimers with ErbB2.  Specific C-terminal tyrosine residues are then autophosphorylated and, in turn, bind to adaptor proteins, kinases, or protein tyrosine phosphatases.  In addition, c-Src-dependent phosphorylations at other sites, such as tyrosine 845 (Y845) in the kinase domain, regulate the receptor's kinase activity.  Inappropriate expression or mutations of EGFR and/or deregulation of its signaling pathways are associated with many types of cancer, making EGFR a promising target for cancer therapies.

      The 12A3 monoclonal antibody recognizes the phosphorylated Y845 in the protein tyrosine kinase domain of human EGFR.

      558523 Rev. 4
      Format Details
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      Alexa Fluor™ 647
      Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor™ 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 670-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
      Alexa Fluor™ 647
      Red 627-640 nm
      653 nm
      669 nm
      558523 Rev.4
      Citations & References
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      View product citations for antibody "558523" on CiteAb

      Development References (7)

      1. Biscardi JS, Maa ME, Tice DA, Cox ME, Leu TH, Parsons SJ. c-Src-mediated phosphorylation of the epidermal growth factor receptor on Tyr845 and Tyr1101 is associated with modulation of receptor function. J Biol Chem. 1999; 274(12):8335-8343. (Biology).
      2. Boerner JL, Demory ML, Silva C, Parsons SJ. Phosphorylation of Y845 on the epidermal growth factor receptor mediates binding to the mitochondrial protein cytochrome c oxidase subunit II. Mol Cell Biol. 2004; 24(16):7059-7071. (Biology).
      3. Hristozova T, Konschak R, Budach V, Tinhofer I. A simple multicolor flow cytometry protocol for detection and molecular characterization of circulating tumor cells in epithelial cancers. Cytometry A. 2012; 81A(6):489-495. (Clone-specific: Flow cytometry). View Reference
      4. Mendelsohn J, Baselga J. Status of epidermal growth factor receptor antagonists in the biology and treatment of cancer. J Clin Oncol. 2003; 21(14):2787-2799. (Biology).
      5. Moro L, Dolce L, Cabodi S, et al. Integrin-induced epidermal growth factor (EGF) receptor activation requires c-Src and p13Cas and leads to phosphorylation of specific EGF receptor tyrosines. J Biol Chem. 2002; 277(11):9405-9414. (Biology).
      6. Olayioye MA, Neve RM, Lane HA, Nynes NE. The ErbB signaling network: receptor heterodimerization in development and cancer. EMBO J. 2000; 19(13):3159-3167. (Biology).
      7. Wu W, Graves LM, Gill GN, Parsons SJ, Samet JM. Src-dependent phosphorylation of the epidermal growth factor receptor on tyrosine 845 is required for zinc-induced Ras activation. J Biol Chem. 2002; 277(27):24252-24257. (Biology).
      View All (7) View Less
      558523 Rev. 4

       

      Please refer to Support Documents for Quality Certificates


      Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


      Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

      For Research Use Only. Not for use in diagnostic or therapeutic procedures.