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      Alexa Fluor® 647 Mouse anti-MEK1
      Alexa Fluor® 647 Mouse anti-MEK1
      Analysis of MEK1 in HeLaS3 cells.  HeLaS3 cells were either transfected with MEK1 RNAi (open histogram) or untreated (shaded histogram).  The cells were fixed (BD Cytofix™ Fixation 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-MEK1 (Cat. No. 560101).  Down-regulation of MEK1 expression is evident in the RNAi-transfected cells.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.
      Alexa Fluor® 647 Mouse anti-MEK1
      Analysis of MEK1 in HeLaS3 cells.  HeLaS3 cells were either transfected with MEK1 RNAi (open histogram) or untreated (shaded histogram).  The cells were fixed (BD Cytofix™ Fixation 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-MEK1 (Cat. No. 560101).  Down-regulation of MEK1 expression is evident in the RNAi-transfected cells.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.
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      BD Phosflow™
      MAPK/ERK kinase 1, EC 2.7.12.2, kinase MEK1, MAPKK1, PRKMK1
      Human (QC Testing), Mouse, Rat, Dog, Chicken (Reported)
      Mouse IgG2a
      Human MEK1 Recombinant Protein
      Intracellular staining (flow cytometry) (Routinely Tested)
      20 µl
      AB_1645607
      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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      推奨アッセイ手順

      Either BD Cytofix™ fixation buffer or BD Phosflow™ Fix Buffer I may be used for cell fixation.

      This mAb was characterized by flow cytometry (Flow) and western blot analysis (WB) using these model systems:

      Method                Species                Cells                        Treatment                        Fixation                        Perm buffer                Result

      Flow                        Human                HeLaS3                        RNAi                                BD Cytofix™                 III                                Down-regulation

      Flow                        Human                PBMC                        Not Applicable                BD Cytofix™                 I, II or III                        Positive Staining

      WB                        Human                A431 Cell Lysate        Not Applicable                Not Applicable        Not Applicable        45 kDa

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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. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
      3. 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.
      4. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
      5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      6. 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.
      7. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
      8. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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      560101 Rev. 2
      抗体の詳細
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      25/MEK1

      MEK1 (MapK/ERK Kinase 1) is a 45-kDa member of the MEK family of dual specificity kinases.  MEK is activated by a variety of cellular serine/threonine kinases including c-Raf, A-Raf, c-mos, and MEK Kinase-1.  Activated MEK phosphorylates MAP kinase (ERK) at threonine and tyrosine residues.  This results in activation of ERK and its signaling pathway. MEK is highly specific for ERK and various MEKs preferentially phosphorylate individual ERK isoforms.  MEK1 only activates ERK1 and ERK2.  This specificity may result from variations in ERK regions that are known as the phosphorylation lip and kinase backbone.  MEK's localization is cytoplasmic, but mitogenic stimulation induces a mass translocation to the nucleus.  Mechanisms behind this nuclear translocation remain unknown.  However, MEK contains an N-terminal nuclear export signal (NES) that mediates its rapid exodus from the nucleus and restores its unstimulated cellular distribution.

      The 25/MEK1 monoclonal antibody recognizes MEK1, regardless of phosphorylation status.

      The specificity of this antibody conjugate for flow cytometric analysis was validated by confirming that RNA-mediated interference (RNAi) of the specific protein reduced the staining of the cells (see figure). Furthermore, the capacity of the RNAi to down-regulate the expression of the relevant protein was confirmed by western blot analysis.

      560101 Rev. 2
      フォーマットの詳細
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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 520-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
      560101 Rev.2
      引用＆参考文献
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      Development References (4)

      1. Aplin AE, Stewart SA, Assoian RK, Juliano RL. Integrin-mediated adhesion regulates ERK nuclear translocation and phosphorylation of Elk-1. J Cell Biol. 2001; 153(2):273-282. (Clone-specific: Immunofluorescence, Western blot). View Reference
      2. Gu J, Fujibayashi A, Yamada KM, Sekiguchi K. Laminin-10/11 and fibronectin differentially prevent apoptosis induced by serum removal via phosphatidylinositol 3-kinase/Akt- and MEK1/ERK-dependent pathways. J Biol Chem. 2002; 277(22):19922-19928. (Clone-specific: Western blot). View Reference
      3. Robinson MJ, Cheng M, Khokhlatchev A, et al. Contributions of the mitogen-activated protein (MAP) kinase backbone and phosphorylation loop to MEK specificity. J Biol Chem. 1996; 271(47):29734-29739. (Biology). View Reference
      4. Short SM, Boyer JL, Juliano RL. Integrins regulate the linkage between upstream and downstream events in G protein-coupled receptor signaling to mitogen-activated protein kinase. J Biol Chem. 2000; 275(17):12970-12977. (Clone-specific: Immunoprecipitation, In vitro kinase assay, Western blot). View Reference
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      560101 Rev. 2

      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.

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