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      Alexa Fluor™ 647 Rat Anti-Mouse AIRE
      Alexa Fluor™ 647 Rat Anti-Mouse AIRE
      Analysis of Aire Expression        Panel A. Multicolor flow cytometric analysis of Aire expression in thymic medullary epithelial cells (mTECs) from wild type (Aire +/+) and knockout (Aire –/–) mice. Enriched preparation of mTECs prepared from wild type (Aire +/+; Left 2 Plots) and knockout (Aire –/–; Right 2 Plots) mice were fixed and permeabilized with BD Transcription Factor Buffer Set (Cat. No. 562574/562725). The cells were then stained with fluorescent antibodies specific for CD45, Ly51, and EpCAM, fluorescent Ulex europaeus agglutinin 1 (UEA1), FITC Anti-CD80 and either Alexa Fluor™ 647 Rat IgG2a κ Isotype Control (Cat. No. 557906) or Alexa Fluor™ 647 Rat Anti-Mouse Aire antibody (Cat. No. 567253) at 0.25 μg/test. Aire expression was evaluated after gating for CD45- EpCAM+ Ly51– UEA1+ cells (see reference J Immunol 2018 201:3244 for further details). The bivariate pseudocolor density plots showing the correlated expression of Aire (or Ig Isotype control staining) versus CD80 were derived from gated events with the forward and side light-scatter characteristics of intact mTECs. Flow cytometry and data analysis were performed using a BD FACSAria™ II Cell Analyzer System and BD FACSDiva™ Software. (The flow data figures were kindly provided by Dr. Mitsuru Matsumoto's laboratory).        Panel B. Flow cytometric analysis of Aire expression in mouse Aire-transfected and -untransfected cells. The untransfected control 293F (dotted line histogram) and mouse Aire-transfected 293F (solid line histogram) cells were fixed and permeabilized with BD Transcription Factor Buffer Set. The cells were then stained with Alexa Fluor™ 647 Rat Anti-Mouse Aire antibody at 0.5 μg/test. Flow cytometric analysis was performed using a BD LSRFortessa™ X-20 Flow Cytometer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
      Alexa Fluor™ 647 Rat Anti-Mouse AIRE
      Analysis of Aire Expression        Panel A. Multicolor flow cytometric analysis of Aire expression in thymic medullary epithelial cells (mTECs) from wild type (Aire +/+) and knockout (Aire –/–) mice. Enriched preparation of mTECs prepared from wild type (Aire +/+; Left 2 Plots) and knockout (Aire –/–; Right 2 Plots) mice were fixed and permeabilized with BD Transcription Factor Buffer Set (Cat. No. 562574/562725). The cells were then stained with fluorescent antibodies specific for CD45, Ly51, and EpCAM, fluorescent Ulex europaeus agglutinin 1 (UEA1), FITC Anti-CD80 and either Alexa Fluor™ 647 Rat IgG2a κ Isotype Control (Cat. No. 557906) or Alexa Fluor™ 647 Rat Anti-Mouse Aire antibody (Cat. No. 567253) at 0.25 μg/test. Aire expression was evaluated after gating for CD45- EpCAM+ Ly51– UEA1+ cells (see reference J Immunol 2018 201:3244 for further details). The bivariate pseudocolor density plots showing the correlated expression of Aire (or Ig Isotype control staining) versus CD80 were derived from gated events with the forward and side light-scatter characteristics of intact mTECs. Flow cytometry and data analysis were performed using a BD FACSAria™ II Cell Analyzer System and BD FACSDiva™ Software. (The flow data figures were kindly provided by Dr. Mitsuru Matsumoto's laboratory).        Panel B. Flow cytometric analysis of Aire expression in mouse Aire-transfected and -untransfected cells. The untransfected control 293F (dotted line histogram) and mouse Aire-transfected 293F (solid line histogram) cells were fixed and permeabilized with BD Transcription Factor Buffer Set. The cells were then stained with Alexa Fluor™ 647 Rat Anti-Mouse Aire antibody at 0.5 μg/test. Flow cytometric analysis was performed using a BD LSRFortessa™ X-20 Flow Cytometer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.
      Product Details
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      BD Pharmingen™
      APECED protein homolog; autoimmune polyendocrinopathy candidiasis ectodermal dystrophy protein homolog; autoimmune regulator
      Mouse (QC Testing)
      Rat IgG2a, κ
      Mouse Aire peptide
      Intracellular staining (flow cytometry) (Routinely Tested)
      0.2 mg/ml
      Aqueous buffered solution containing ≤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 the dye under optimum conditions and unreacted dye was removed.
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      Recommended Assay Procedures

      BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation).  When fluorochrome conjugated antibodies are bound to CompBeads, they have spectral properties very similar to cells.   However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls.  It is strongly recommended that when using a reagent for the first time, users compare the spillover on cell and CompBead to ensure that BD Comp beads are appropriate for your specific cellular application.

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      Product Notices

      1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
      2. An isotype control should be used at the same concentration as the antibody of interest.
      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. This product is provided under an intellectual property license between Life Technologies Corporation and BD Businesses. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The buyer cannot sell or otherwise transfer (a) this product (b) its components or (c) materials made using this product or its components to a third party or otherwise use this product or its components or materials made using this product or its components for Commercial Purposes. Commercial Purposes means any activity by a party for consideration and may include, but is not limited to: (1) use of the product or its components in manufacturing; (2) use of the product or its components to provide a service, information, or data; (3) use of the product or its components for therapeutic, diagnostic or prophylactic purposes; or (4) resale of the product or its components, whether or not such product or its components are resold for use in research. For information on purchasing a license to this product for any other use, contact Life Technologies Corporation, Cell Analysis Business Unit Business Development, 29851 Willow Creek Road, Eugene, OR 97402, USA, Tel: (541) 465-8300. Fax: (541) 335-0504.
      7. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
      8. Alexa Fluor™ is a trademark of Life Technologies Corporation.
      9. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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      567253 Rev. 1
      Antibody Details
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      MM-525

      The MM-525 monoclonal antibody specifically recognizes mouse Aire (Autoimmune Regulator). This transcription factor is mainly expressed in the thymus, the primary lymphoid organ for T cell development. Aire expression is restricted to medullary thymic epithelial cells (mTECs). Aire induces the expression of a large variety of organ-specific self-antigens that are normally expressed in the peripheral tissues. mTECs participate in the negative selection of developing autoreactive T cells and the induction of regulatory T cells by presenting them with self-antigens in the context of MHC class I and class II molecules. In this way, mTECs play a vital role as self-antigen presenting cells in establishing immunological self-tolerance. A defect in the human AIRE gene causes a rare autosomal recessive systemic autoimmune disease called autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED). The MM-525 monoclonal antibody is useful for detecting Aire expression in mouse tissues, including medullary thymic epithelial cells.

      567253 Rev. 1
      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 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
      567253 Rev.1
      Citations & References
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      View product citations for antibody "567253" on CiteAb

      Development References (2)

      1. Morimoto, J., Y. Nishikawa, et al. Aire Controls in Trans the Production of Medullary Thymic Epithelial Cells Expressing Ly-6C/Ly-6G.. J Immunol. 2018; 201:3244-3257. (Biology). View Reference
      2. Zhao, B., L. Chang, H. Fu, G. Sun, and W. Yang. The Role of Autoimmune Regulator (AIRE) in Peripheral Tolerance.. J Immunol Res. 2018; 3930750 . (Biology). View Reference
      567253 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.