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      PE-CF594 Mouse Anti-ERK1/2 (pT202/pY204)
      PE-CF594 Mouse Anti-ERK1/2 (pT202/pY204)
      Analysis of Erk1/2 (pT202/pY204) in human peripheral blood lymphocytes. Whole blood was either left untreated (dashed line histogram) or treated (solid line histogram) with 400 nM Phorbol 12-Myristate 13-Acetate (PMA; Sigma, Cat. No. P8139) for 15 minutes at 37°C. Erythrocytes were lysed and the leucocytes were fixed with 1X BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10 minutes at 37°C. The cells were then permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes and stained with BD Horizon™ PE-CF594 anti-Erk1/2 (pT202/pY204) (Cat No. 562644). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
      PE-CF594 Mouse Anti-ERK1/2 (pT202/pY204)
      Analysis of Erk1/2 (pT202/pY204) in human peripheral blood lymphocytes. Whole blood was either left untreated (dashed line histogram) or treated (solid line histogram) with 400 nM Phorbol 12-Myristate 13-Acetate (PMA; Sigma, Cat. No. P8139) for 15 minutes at 37°C. Erythrocytes were lysed and the leucocytes were fixed with 1X BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) for 10 minutes at 37°C. The cells were then permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice for 30 minutes and stained with BD Horizon™ PE-CF594 anti-Erk1/2 (pT202/pY204) (Cat No. 562644). The fluorescence histograms were derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes. Flow cytometry was performed using a BD™ LSR II Flow Cytometer System.
      Product Details
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      BD Phosflow™
      p44/42 MAPK; Extracellular signal-Regulated Kinase 1/2 (pT202/Y204)
      Human (QC Testing)
      Mouse IgG1
      Phosphorylated Rat ERK1 (T202/Y204) Peptide
      Intracellular staining (flow cytometry) (Routinely Tested)
      5 µl
      AB_2737698
      Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
      RUO


      Preparation And Storage

      The antibody was conjugated with BD Horizon™ PE-CF594 under optimum conditions, and unconjugated antibody and free PE-CF594 were removed. 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.
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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. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      4. Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
      5. 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.
      6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      7. Texas Red is a registered trademark of Molecular Probes, Inc., Eugene, OR.
      8. CF™ is a trademark of Biotium, Inc.
      9. When excited by the yellow-green (561-nm) laser, the fluorescence may be brighter than when excited by the blue (488-nm) laser.
      10. This product is provided under an Agreement between BIOTIUM and BD Biosciences. The manufacture, use, sale, offer for sale, or import of this product is subject to one or more patents or pending applications owned or licensed by Biotium, Inc. This product, and only in the amount purchased by buyer, may be used solely for buyer’s own internal research, in a manner consistent with the accompanying product literature. No other right to use, sell or otherwise transfer (a) this product, or (b) its components is hereby granted expressly, by implication or by estoppel. This product is for research use only. Diagnostic uses require a separate license from Biotium, Inc. For information on purchasing a license to this product including for purposes other than research, contact Biotium, Inc., 3159 Corporate Place, Hayward, CA 94545, Tel: (510) 265-1027. Fax: (510) 265-1352. Email: btinfo@biotium.com.
      11. Because of the broad absorption spectrum of the tandem fluorochrome, extra care must be taken when using multi-laser cytometers, which may directly excite both PE and CF™594.
      12. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
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      562644 Rev. 1
      Antibody Details
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      20A

      The members of the Mitogen-Activated Protein Kinase (MAPK) family are components of a key signal transduction cascade that links events at the cell surface to responses in the nucleus. The signaling cascade is found in species as varied as yeast and humans, with many of the proteins being well conserved. In mammals the most widely studied members of the cascade are the Extracellular signal-Regulated Kinases, ERK1 (p44 MAPK) and ERK2 (p42 MAPK).  ERK1 and ERK2 share 85% homology and are activated by extracellular signals such as growth factors, hormones, and phorbol esters. Activation occurs through a series of phosphorylations by kinases activating other kinases and eventually leading to phosphorylation of the ERKs. Growth factor stimulation leads to activation of Ras and Raf, leading to phosphorylation of MEK1 (MAPK/ERK kinase) which, in turn, activates the ERKs via dual phosphorylation. Once activated, the ERKs phosphorylate other cytoplasmic signalling molecules, cell-surface receptors, microtubule-associated proteins, and transcription factors in the nucleus. Thus, the active ERK has myriad downstream effectors that implicate it in the control of cell proliferation and differentiation, as well as regulation of the cytoskeleton. Furthermore, studies have shown that elevated ERK activity is associated with some cancers.

      The 20A monoclonal antibody recognizes the phosphorylated threonine 202 and tyrosine 204 (pT202/pY204) of human ERK1 and pT184/pY186 of human ERK2. The orthologous phosphorylation sites in murine ERK1 and ERK2 are T203/Y205 and T183/Y185.

      This antibody is conjugated to BD Horizon™ PE-CF594, which has been developed exclusively by BD Biosciences as a better alternative to PE-Texas Red®. PE-CF594 excites and emits at similar wavelengths to PE-Texas Red® yet exhibits improved brightness and spectral characteristics. Due to PE having maximal absorption peaks at 496 nm and 564 nm, PE-CF594 can be excited by the blue (488-nm), green (532-nm) and yellow-green (561-nm) lasers and can be detected with the same filter set as PE-Texas Red® (eg 610/20-nm filter).

      562644 Rev. 1
      Format Details
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      PE-CF594
      BD Horizon™ PE-CF594 dye is a part of the BD PE family of dyes. This tandem fluorochrome is comprised of a R-Phycoerythrin (PE) donor that has excitation maxima (Ex Max) of 496-nm and 566-nm and an acceptor dye with an emission maximum (Em Max) at 615-nm. PE-CF594, driven by BD innovation, is designed to be excited by the blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and detected using an optical filter centered near 615 nm (e.g., a 610/20-nm bandpass filter). The donor dye can be excited by the Blue (488-nm), Green (532-nm) and yellow-green (561-nm) lasers and the acceptor dye can be excited by the green (532-nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
      altImg
      PE-CF594
      Yellow-Green 488 nm, 532 nm, 561 nm
      496 nm, 566 nm
      615 nm
      562644 Rev.1
      Citations & References
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      Development References (3)

      1. Boulton TG, Cobb MH. Identification of multiple extracellular signal-regulated kinases (ERKs) with antipeptide antibodies. Cell Regul. 1991; 2(5):357-371. (Biology). View Reference
      2. Clark EA, Hynes RO. Ras activation is necessary for integrin-mediated activation of extracellular signal-regulated kinase 2 and cytosolic phospholipase A2 but not for cytoskeletal organization. J Biol Chem. 1996; 271(25):14814-14818. (Biology). View Reference
      3. Sivaraman VS, Wang H, Nuovo GJ, Malbon CC. Hyperexpression of mitogen-activated protein kinase in human breast cancer. J Clin Invest. 1997; 99(7):1478-1483. (Biology). View Reference
      562644 Rev. 1

      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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