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FITC Mouse Anti-Human Granzyme A Reagent Set

FITC Mouse Anti-Human Granzyme A Reagent Set

(RUO)
FITC Mouse Anti-Human Granzyme A Reagent Set

Profile of permeabilized peripheral blood lymphocytes analyzed by flow cytometry (BDIS, San Jose, CA). Cells were collected, fixed, and permeabilized using the Cytofix/Cytoperm™ Kit (Cat. No. 554715) for 20 minutes at room temperature (RT), pelleted and washed twice with Perm/Wash Buffer™. Cells were resuspended in Perm/Wash Buffer and stained with FITC anti-human Granzyme A (clone CB9, Cat. No. 558905) 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.

Profile of permeabilized peripheral blood lymphocytes analyzed by flow cytometry (BDIS, San Jose, CA). Cells were collected, fixed, and permeabilized using the Cytofix/Cytoperm™ Kit (Cat. No. 554715) for 20 minutes at room temperature (RT), pelleted and washed twice with Perm/Wash Buffer™. Cells were resuspended in Perm/Wash Buffer and stained with FITC anti-human Granzyme A (clone CB9, Cat. No. 558905) 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)
Flow cytometry (Routinely Tested)
RUO
AB_397156


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). The physiological substrates for granzyme A in the apoptotic pathway have not been identified. 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.  

Clone CB9 recognizes human granzyme A. Purified human granzyme A was used as immunogen. Clone MOPC-21 is a mouse IgG1 isotype (negative) control. The MOPC-21 antibody has unknown specificity.  CB9 and MOPC-21 FITC conjugates are optimized in F/P ratios.

The antibodies are routinely tested in parallel by flow cytometry. Other applications were tested at BD Biosciences Pharmingen during antibody development only or reported in the literature.

Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with FITC under optimum conditions, and unreacted FITC was removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.

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. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  3. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  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.
  5. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
558905 Rev. 5
Components
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Description Quantity/Size Part Number EntrezGene ID
FITC Human Granzyme A 100 Tests (1 ea) 51-68394X N/A
FITC Mouse IgG1, κ Isotype Control 100 Tests (1 ea) 51-13854X-8 N/A
558905 Rev. 5
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: Immunoprecipitation). 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
558905 Rev. 5

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.

Refer to manufacturer's instructions for use and related User Manuals and Technical Data Sheets before using this product 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.