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      PE Mouse Anti-Human Granzyme A Set

      BD Pharmingen™ PE Mouse Anti-Human Granzyme A Set

      (RUO)
      PE Mouse Anti-Human Granzyme A Set
      Flow cytometric analysis of Granzyme A expression in human peripheral blood lymphocytes. Human peripheral blood mononuclear cells (PBMC) were fixed using BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722). The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either PE Mouse IgG1 Isotype Control (Component No. 51-13855X-6; dashed line histogram) or PE Mouse Anti-Human Granzyme A antibody (Component No. 51-68395X; solid line histogram). The fluorescence histogram showing the expression of Granzyme A (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes.
      PE Mouse Anti-Human Granzyme A Set
      Profile of permeabilized peripheral blood lymphocytes analyzed by flow cytometry (BDIS, San Jose, CA). Cells were collected, fixed, and permeabilzied using the Cytofix/Cytoperm™ Kit (554714) for 20 minutes at room temperature (RT), pelleted, and washed twice with Perm/Wash Buffer™ (component of Cat. No. 554714). Cells were resuspended in Perm/Wash Buffer™ and stained with PE Anti-human Granzyme A (Clone CB9) or with an isotype control (clone MOPC-21) for 20-30 minutes at RT in the dark. Cells were then washed once in Perm/Wash Buffer™, resuspended in wash buffer, and analyzed by flow cytometry.
      PE Mouse Anti-Human Granzyme A Set PE Mouse Anti-Human Granzyme A Set PE Mouse Anti-Human Granzyme A Set
      Flow cytometric analysis of Granzyme A expression in human peripheral blood lymphocytes. Human peripheral blood mononuclear cells (PBMC) were fixed using BD Cytofix/Cytoperm™ Fixation and Permeabilization Solution (Cat. No. 554722). The cells were then washed and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either PE Mouse IgG1 Isotype Control (Component No. 51-13855X-6; dashed line histogram) or PE Mouse Anti-Human Granzyme A antibody (Component No. 51-68395X; solid line histogram). The fluorescence histogram showing the expression of Granzyme A (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of intact lymphocytes.
      Profile of permeabilized peripheral blood lymphocytes analyzed by flow cytometry (BDIS, San Jose, CA). Cells were collected, fixed, and permeabilzied using the Cytofix/Cytoperm™ Kit (554714) for 20 minutes at room temperature (RT), pelleted, and washed twice with Perm/Wash Buffer™ (component of Cat. No. 554714). Cells were resuspended in Perm/Wash Buffer™ and stained with PE Anti-human Granzyme A (Clone CB9) or with an isotype control (clone MOPC-21) for 20-30 minutes at RT in the dark. Cells were then washed once in Perm/Wash Buffer™, resuspended in wash buffer, and analyzed by flow cytometry.
      Product Details
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      BD Pharmingen™
      Human (QC Testing)
      Intracellular staining (flow cytometry) (Routinely Tested)
      RUO
      AB_397155


      Description

      The primary mechanism by which cytotoxic T cells eliminate virally infected cells is by granule exocytosis. The release of cytotoxic granule contents by cytotoxic T lymphocytes (CTL) triggers apoptotic target cell death. CTL granules contain a pore-forming protein, perforin, and a group of serine proteases called granzymes. In the classic model, perforins create holes in the target cell membrane, allowing entrance of the granzymes. Granzyme A and B are the predominant granzymes activated after CTL activation, but each act via an independent apoptotic pathway; granzyme B is activated immediately, while granzyme A acts hours later. Granzyme B does not induce cleavage of caspase-3, lamin B, rho-GTPase or PARP, but does cleave DNA-PKcs and nuclear mitotic apparatus protein (NuMA). Studies involving mice which are deficient in both granzyme A and B suggest a model whereby the granzyme B pathway may have evolved as the major apoptotic pathway with the granzyme A pathway acting as a backup. However, further research is needed to delineate the components of the distinct pathways.

      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 unconjugated antibody and free dye were 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 cells and BD CompBeads to ensure that BD™ CompBeads are appropriate for your specific cellular application.

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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. 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. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
      5. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
      6. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
      7. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
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      558904 Rev. 6
      Components
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      Description Quantity/Size Part Number EntrezGene ID
      PE Granzyme A 100 Tests (1 ea) 51-68395X N/A
      PE Mouse IgG1, κ Isotype Control 100 Tests (1 ea) 51-13855X-6 N/A
      558904 Rev. 6
      Citations & References
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      Development References (5)

      1. Andrade F, Roy S, Nicholson D, Thornberry N, Rosen A, Casciola-Rosen L. Granzyme B directly and efficiently cleaves several downstream caspase substrates: implications for CTL-induced apoptosis. Immunity. 1998; 8(4):451-460. (Biology). View Reference
      2. Beresford PJ, Kam CM, Powers JC, Lieberman J. Recombinant human granzyme A binds to two putative HLA-associated proteins and cleaves one of them. Proc Natl Acad Sci U S A. 1997; 94(17):9285-9290. (Immunogen: Immunoprecipitation). View Reference
      3. Beresford PJ, Xia Z, Greenberg AH, Lieberman J. Granzyme A loading induces rapid cytolysis and a novel form of DNA damage independently of caspase activation. Immunity. 1999; 10(5):585-594. (Clone-specific). View Reference
      4. Shresta S, Graubert TA, Thomas DA, Raptis SZ, Ley TJ. Granzyme A initiates an alternative pathway for granule-mediated apoptosis. Immunity. 1999; 10(5):595-605. (Biology). View Reference
      5. Trimble LA, Lieberman J. Circulating CD8 T lymphocytes in human immunodeficiency virus-infected individuals have impaired function and downmodulate CD3 zeta, the signaling chain of the T-cell receptor complex. Blood. 1998; 91(2):585-594. (Clone-specific: Flow cytometry). View Reference
      558904 Rev. 6

      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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