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Alexa Fluor® 647 Mouse anti-SLP-76 (pY128)
Alexa Fluor® 647 Mouse anti-SLP-76 (pY128)
Analysis of SLP-76 (pY128) in human peripheral blood lymphocytes.  Human peripheral blood mononuclear cells were either unstimulated (open histograms) or stimulated by cross-linking with CD3 (Purified NA/LE Mouse anti-Human CD3 mAb, clone UCHT1,Cat. No. 555329) and CD28 (Purified NA/LE Mouse anti-Human CD28, clone CD28.2, Cat. No. 555725) on ice for 15 minutes followed by incubation with the Purified Goat anti-Mouse Ig (Cat. No. 553998) on ice for 15 minutes, and then allowed to undergo phosphorylation at 37°C for 5 minutes (shaded histograms). The cells were fixed (BD Cytofix™ Fixation buffer Cat. No. 554655) for 10 minutes at 37°C, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, blocked with normal mouse immunoglobulin, and then stained with Alexa Fluor® 647 Mouse anti-SLP-76 (pY128) (Cat. No. 558438) and PerCP-Cy5.5 Mouse anti-human CD3 mAb SK7 (Cat. No. 340949).  The left panel displays the upregulated phosphorylation of SLP-76 in activated CD3-positive T lymphocytes, while the right panel displays the lack of upregulated SLP-76 phosphorylation in CD3-negative non-T cells.  Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
Analysis of SLP-76 (pY128) in human peripheral blood lymphocytes.  Human peripheral blood mononuclear cells were either unstimulated (open histograms) or stimulated by cross-linking with CD3 (Purified NA/LE Mouse anti-Human CD3 mAb, clone UCHT1,Cat. No. 555329) and CD28 (Purified NA/LE Mouse anti-Human CD28, clone CD28.2, Cat. No. 555725) on ice for 15 minutes followed by incubation with the Purified Goat anti-Mouse Ig (Cat. No. 553998) on ice for 15 minutes, and then allowed to undergo phosphorylation at 37°C for 5 minutes (shaded histograms). The cells were fixed (BD Cytofix™ Fixation buffer Cat. No. 554655) for 10 minutes at 37°C, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes, blocked with normal mouse immunoglobulin, and then stained with Alexa Fluor® 647 Mouse anti-SLP-76 (pY128) (Cat. No. 558438) and PerCP-Cy5.5 Mouse anti-human CD3 mAb SK7 (Cat. No. 340949).  The left panel displays the upregulated phosphorylation of SLP-76 in activated CD3-positive T lymphocytes, while the right panel displays the lack of upregulated SLP-76 phosphorylation in CD3-negative non-T cells.  Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
製品詳細
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BD Phosflow™
Human (QC Testing), Mouse,Rat (Predicted)
Mouse BALB/c IgG1, κ
Phosphorylated Human SLP-76
Intracellular staining (flow cytometry) (Routinely Tested)
20 µl
AB_647159
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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This antibody conjugate is suitable for intracellular staining of human whole blood (using BD Phosflow™ Lyse/Fix Buffer, Cat. No. 558049) and peripheral blood mononuclear cells (using BD Cytofix™ Fixation Buffer, Cat. No. 554655). Any of the 3 BD Phosflow™ Perm Buffers may be used.

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. 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.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. 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.
  6. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
  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.
558438 Rev. 3
抗体の詳細
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J141-668.36.58

SLP-76 (SH2 domain-containing Leukocyte Protein of 76 kDa) is a tyrosine phosphoprotein that is involved in the T cell receptor (TCR)-mediated intracellular signaling pathway.  It may be involved in the signaling pathways of other peripheral blood leukocytes; thymic/splenic cells; and in human T, B, and monocytic cell lines.  SLP-76 consists of several motifs that signify its importance in protein-protein interactions involved in intracellular signaling pathways, such as the SH2 domain in the C-terminus, the three amino-terminus 17-amino acid repeats with conserved tyrosine and acidic residues (DYE(S/P)P), and a proline rich region.  SLP-76 has been shown to associate with Gads, Grb2, PLCγ1, SLAP-130, and Vav, all of which are part of the signaling cascade in T lymphocytes.  An early event in the T cell activation pathway is the phosphorylation, by the Syk-family kinase ZAP-70, of SLP-76 at the three conserved tyrosine motifs, which then mediate interactions with downstream effectors.  The phosphorylated tyrosine 128 (Y128) brings the Rho family guanine nucleotide exchange factor Vav1 and the Nck adapter protein, which binds to p21-activated kinase (PAK1) and Wiskott-Aldrich syndrome protein (WASP), into the activation complex.  Vav1, PAK1, and WASP may mediate TCR-stimulated actin cytoskeletal rearrangement.  

The J141-688.36.58 monoclonal antibody recognizes the phosphorylated Y128 of activated SLP-76.

558438 Rev. 3
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 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
558438 Rev.3
引用&参考文献
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View product citations for antibody "558438" on CiteAb

Development References (3)

  1. Fang N, Motto DG, Ross SE, Koretzky GA. Tyrosines 113, 128, and 145 of SLP-76 are required for optimal augmentation of NFAT promoter activity. J Immunol. 1996; 157:3769-3773. (Biology). View Reference
  2. Janssen E, Zhang W. Adaptor proteins in lymphocyte activation. Curr Opin Immunol. 2003; 15:269-276. (Biology). View Reference
  3. Wu JN, Koretzky GA. The SLP-76 family of adapter proteins. Semin Immunol. 2004; 16:379-393. (Biology). View Reference
558438 Rev. 3

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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.