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      PE Mouse Anti-Human RANTES
      PE Mouse Anti-Human RANTES
      Expression of RANTES by cultured human PBMC. Human PBMC were cultured for 24 hours in the presence of GolgiStop™ (2 µM final concentration; Cat. No. 554724). The PBMC were harvested, stained with FITC-mouse anti-human CD3 antibody (FITC-HIT3a, Cat. No. 555339), fixed, permeabilized, and subsequently stained with 0.125 µg of PE-mouse anti-human RANTES antibody (PE-2D5, Cat. No. 554732) following Pharmingen's staining protocol (left panel). To demonstrate specificity of staining, the binding of PE-2D5 antibody was blocked by the preincubation of the unlabeled 2D5 antibody with fixed/permeabilized target cells prior to staining with the PE-2D5 antibody. The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with unlabeled antibody blocking control (right panel).
      PE Mouse Anti-Human RANTES
      Expression of RANTES by cultured human PBMC. Human PBMC were cultured for 24 hours in the presence of GolgiStop™ (2 µM final concentration; Cat. No. 554724). The PBMC were harvested, stained with FITC-mouse anti-human CD3 antibody (FITC-HIT3a, Cat. No. 555339), fixed, permeabilized, and subsequently stained with 0.125 µg of PE-mouse anti-human RANTES antibody (PE-2D5, Cat. No. 554732) following Pharmingen's staining protocol (left panel). To demonstrate specificity of staining, the binding of PE-2D5 antibody was blocked by the preincubation of the unlabeled 2D5 antibody with fixed/permeabilized target cells prior to staining with the PE-2D5 antibody. The quadrant markers for the bivariate dot plots were set based on the autofluorescence control, and verified with unlabeled antibody blocking control (right panel).
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      BD Pharmingen™
      Human (QC Testing)
      Mouse BALB/c IgG1, κ
      Recombinant Human RANTES Protein
      Intracellular staining (flow cytometry) (Routinely Tested)
      0.2 mg/ml
      AB_395534
      Aqueous buffered solution containing ≤0.09% sodium azide.
      RUO


      Preparation and Storage

      The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.
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      推奨アッセイ手順

      Immunofluorescent Staining and Flow Cytometry: The PE-conjugated 2D5 antibody (Cat. No. 554732) can be used for multicolor immunofluorescent staining and flow cytometric analyses to identify and enumerate human RANTES producing cells within mixed cell populations. For optimal immunofluorescent staining with flow cytometric analysis, this anti-cytokine antibody should be titrated (≤ 0.5 µg mAb/million cells). For specific methodology, visit the protocols section of our website, or the chapter on intracellular staining in the Immune Function Handbook, which is posted on our web site at www.bdbiosciences.com.

      A useful control for demonstrating specificity of staining is to pre-block the fixed/permeabilized cells with unlabeled 2D5 antibody (Cat. No. 556859) prior to staining. The intracellular staining technique and use of blocking controls are described in detail by C. Prussin and D. Metcalfe. A suitable mouse IgG1 isotype control immunoglobulin for assessing the level of background staining on paraformaldehydefixed/saponin-permeabilized human cells is the PE-MOPC-21 immunoglobulin (Cat. No. 554680); use at comparable concentrations to antibody of interest (e.g., ≤ 0.5 µg mAb/1 million cells).

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      Product Notices

      1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
      2. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      3. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      4. 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.
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      554732 Rev. 2
      抗体の詳細
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      2D5

      The 2D5 monoclonal antibody specifically binds to human Regulated upon Activation, Normal T cells Expressed and Secreted (RANTES). RANTES belongs to the C-C family of chemokines and is also known as CCL5. RANTES is produced by activated macrophages, CD8+ T cells, platelets, fibroblasts, epithelial cells and some tumor cells. RANTES is a chemoattractant and response modifier for a variety of cells including memory T cells, NK cells, dendritic cells, monocytes, eosinophils, basophils, and mast cells. RANTES binds to and transduces intracellular signals through several cell surface chemokine receptors including CCR1, CCR3, CCR4, and CCR5. RANTES can reportedly inhibit the interaction between certain HIV viruses and CCR5 and thus suppress viral infection in vitro.

      554732 Rev. 2
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      PE
      R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE 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 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
      altImg
      PE
      Yellow-Green 488 nm, 532 nm, 561 nm
      496 nm, 566 nm
      576 nm
      554732 Rev.2
      引用＆参考文献
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      Development References (3)

      1. Leonard EJ, Sylvester I, Yoshimura T, et al. Current Protocols in Immunology. John Wiley and Sons; 1995:6.12.1-6.12.26.
      2. Prussin C, Metcalfe DD. Detection of intracytoplasmic cytokine using flow cytometry and directly conjugated anti-cytokine antibodies. J Immunol Methods. 1995; 188(1):117-128. (Methodology: IC/FCM Block). View Reference
      3. Sticherling M, Küpper M, Koltrowitz F, et al. Detection of the chemokine RANTES in cytokine-stimulated human dermal fibroblasts.. J Invest Dermatol. 1995; 105(4):585-91. (Clone-specific). View Reference
      554732 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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