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      Alexa Fluor™ 647 Mouse Anti-Human Neutrophil Elastase
      Alexa Fluor™ 647 Mouse Anti-Human Neutrophil Elastase

      Flow cytometric analysis of Human Neutrophil Elastase expression in Human peripheral blood granulocytes and tumor cell lines.  Human peripheral blood granulocytes (Left Plot) prepared by density gradient centrifugation and cells from the Human Jurkat (Acute T cell leukemia, ATCC® TIB-152™; Middle Plot) and Human U-937 (Histiocytic Lymphoma, ATCC® CRL-1593.2™, Right Plot) cell lines were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), washed and then permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained in BD Perm/Wash™ Buffer with either Alexa Fluor™ 647 Mouse IgG1, κ Isotype Control (Cat. No. 565571; dashed line histograms) or Alexa Fluor™ 647 Mouse Anti-Human Neutrophil Elastase antibody (Cat. No. 569516/569517; solid line histograms) at 1 µg/test. The fluorescence histograms showing Neutrophil Elastase expression (or Ig Isotype control staining) were derived from gated events with the light-scatter characteristics of intact cells. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

      Alexa Fluor™ 647 Mouse Anti-Human Neutrophil Elastase

      Flow cytometric analysis of Human Neutrophil Elastase expression in Human peripheral blood granulocytes and tumor cell lines.  Human peripheral blood granulocytes (Left Plot) prepared by density gradient centrifugation and cells from the Human Jurkat (Acute T cell leukemia, ATCC® TIB-152™; Middle Plot) and Human U-937 (Histiocytic Lymphoma, ATCC® CRL-1593.2™, Right Plot) cell lines were fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), washed and then permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The cells were then stained in BD Perm/Wash™ Buffer with either Alexa Fluor™ 647 Mouse IgG1, κ Isotype Control (Cat. No. 565571; dashed line histograms) or Alexa Fluor™ 647 Mouse Anti-Human Neutrophil Elastase antibody (Cat. No. 569516/569517; solid line histograms) at 1 µg/test. The fluorescence histograms showing Neutrophil Elastase expression (or Ig Isotype control staining) were derived from gated events with the light-scatter characteristics of intact cells. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X20 Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

      Product Details
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      BD Pharmingen™
      ELANE; HNE; PMN elastase; PMN-E; neutrophil elastase
      Human (QC Testing)
      Mouse IgG1
      Neutrophil granule proteins
      Intracellular staining (flow cytometry) (Routinely Tested)
      0.2 mg/ml
      1991
      AB_3685124
      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 BD® 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 BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

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      Product Notices

      1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      2. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
      3. An isotype control should be used at the same concentration as the antibody of interest.
      4. Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
      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. 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.
      8. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
      9. Alexa Fluor™ is a trademark of Life Technologies Corporation.
      10. For U.S. patents that may apply, see bd.com/patents.
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      569516 Rev. 2
      Antibody Details
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      NP57

      The NP57 monoclonal antibody specifically recognizes human Neutrophil Elastase (NE) which is also known as Human leukocyte elastase (HLE), Granulocyte-derived Elastase (GE), or PMN Elastase (PMN-E). Neutrophil Elastase is a major serine proteinase encoded by ELANE (Elastase, neutrophil expressed) that belongs to the elastase subfamily within the peptidase S1 family. Neutrophil Elastase is predominantly present in the azurophilic granules (primary granules) of neutrophils and monocytes. When released outside of these effector cells, it can act to hydrolyze a variety of extracellular matrix substrates including elastin, fibronectin, collagen, and proteoglycans. Elastase activity in normally controlled by serine proteinase inhibitors. Neutrophil Elastase can regulate the functions of NK cells, monocytes, and granulocytes, eg, by inhibiting C5a-dependent enzyme release and chemotaxis by neutrophils. It is also capable of killing bacteria such as E. coli and degrading virulence factors from bacteria such as Yersinia, Shigella, and Salmonella. Dysregulated expression or defects in this enzyme are associated with some inflammatory and degenerative diseases including rheumatoid arthritis, emphysema, chronic obstructive pulmonary disease, and cystic fibrosis as well as neutropenias.

      569516 Rev. 2
      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 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
      569516 Rev.2
      Citations & References
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      View product citations for antibody "569516" on CiteAb

      Development References (7)

      1. Belaaouaj A, Kim KS, Shapiro SD. Degradation of outer membrane protein A in Escherichia coli killing by neutrophil elastase.. Science. 2000; 289(5482):1185-8. (Biology). View Reference
      2. Cramer EM, Beesley JE, Pulford KA, Breton-Gorius J, Mason DY. Colocalization of elastase and myeloperoxidase in human blood and bone marrow neutrophils using a monoclonal antibody and immunogold.. Am J Pathol. 1989; 134(6):1275-84. (Clone-specific: Electron microscopy, Immunofluorescence, Immunohistochemistry). View Reference
      3. Davies PL, Maxwell NC, Kotecha S, Spiller OB. Monoclonal anti-neutrophil elastase antibody characterisation: ability to block function, detect free versus serpin-complexed enzyme and stain intracellular granules.. J Immunol Methods. 2008; 336(2):175-82. (Clone-specific: Intracellular Staining/Flow Cytometry). View Reference
      4. Korkmaz B, Attucci S, Jourdan ML, Juliano L, Gauthier F. Inhibition of neutrophil elastase by alpha1-protease inhibitor at the surface of human polymorphonuclear neutrophils.. J Immunol. 2005; 175(5):3329-38. (Biology). View Reference
      5. Pulford KA, Erber WN, Crick JA, et al. Use of monoclonal antibody against human neutrophil elastase in normal and leukaemic myeloid cells.. J Clin Pathol. 1988; 41(8):853-60. (Immunogen: Immunofluorescence, Immunohistochemistry, Immunoprecipitation, Western blot). View Reference
      6. Tralau T, Meyer-Hoffert U, Schröder JM, Wiedow O. Human leukocyte elastase and cathepsin G are specific inhibitors of C5a-dependent neutrophil enzyme release and chemotaxis.. Exp Dermatol. 2004; 13(5):316-25. (Biology). View Reference
      7. Weinrauch Y, Drujan D, Shapiro SD, Weiss J, Zychlinsky A. Neutrophil elastase targets virulence factors of enterobacteria.. Nature. 2002; 417(6884):91-4. (Biology). View Reference
      View All (7) View Less
      569516 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.