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Biotin Rabbit Anti- Active Caspase-3
Biotin Rabbit Anti- Active Caspase-3

Flow cytometric analysis of apoptotic and non-apoptotic populations for active caspase-3.  Jurkat cells (Human T-cell leukemia; ATCC TIB-152) were left untreated (left panel) or treated with 4 µM of camptothecin for 4 hr to induce apoptosis (right panel). Cells were washed once in PBS, then fixed and permeabilized using the BD Cytofix/Cytoperm™ Kit (Cat. No. 554714) for 20 min at room temperature (RT), pelleted and washed with BD Perm/Wash™ buffer (component of Cat. No. 554714). Cells were subsequently stained with the biotin rabbit anti- active caspase-3 antibody (clone C92-605). Cells were then washed and resuspended in BD Perm/Wash™ buffer containing FITC-Avidin (5 ng/1x10e6 cells) [Cat. No. 554057] and incubated for 30 min at RT in the dark. Cells were then washed and resuspended in BD Perm/Wash™ buffer before analyzing by flow cytometry. The results show that untreated cells were primarily negative for active caspase-3 (left panel, M1); whereas more than half of the treated cells were positive for active caspase-3 staining (right panel, M2).

Flow cytometric analysis of apoptotic and non-apoptotic populations for active caspase-3.  Jurkat cells (Human T-cell leukemia; ATCC TIB-152) were left untreated (left panel) or treated with 4 µM of camptothecin for 4 hr to induce apoptosis (right panel). Cells were washed once in PBS, then fixed and permeabilized using the BD Cytofix/Cytoperm™ Kit (Cat. No. 554714) for 20 min at room temperature (RT), pelleted and washed with BD Perm/Wash™ buffer (component of Cat. No. 554714). Cells were subsequently stained with the biotin rabbit anti- active caspase-3 antibody (clone C92-605). Cells were then washed and resuspended in BD Perm/Wash™ buffer containing FITC-Avidin (5 ng/1x10e6 cells) [Cat. No. 554057] and incubated for 30 min at RT in the dark. Cells were then washed and resuspended in BD Perm/Wash™ buffer before analyzing by flow cytometry. The results show that untreated cells were primarily negative for active caspase-3 (left panel, M1); whereas more than half of the treated cells were positive for active caspase-3 staining (right panel, M2).

Product Details
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BD Pharmingen™
CPP32; Yama; Apopain
Human (QC Testing), Mouse (Tested in Development)
Rabbit IgG
Human Active Caspase-3 Fragment
Intracellular staining (flow cytometry) (Routinely Tested)
20 µl
AB_393750
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


Preparation And Storage

The antibody was conjugated with biotin under optimum conditions, and unreacted biotin was removed. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. 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. 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.
  5. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
550557 Rev. 4
Antibody Details
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C92-605

The caspase family of cysteine proteases plays a key role in apoptosis and inflammation. Caspase-3 is a key protease that is activated during the early stages of apoptosis and, like other members of the caspase family, is synthesized as an inactive pro-enzyme that is processed in cells undergoing apoptosis by self-proteolysis and/or cleavage by another protease. The processed forms of caspases consist of large (17-22 kDa) and small (10-12 kDa) subunits which associate to form an active enzyme. Active caspase-3, a marker for cells undergoing apoptosis, consists of a heterodimer of 17 and 12 kDa subunits which is derived from the 32 kDa pro-enzyme. Active caspase-3 proteolytically cleaves and activates other caspases, as well as relevant targets in the cytoplasm, e.g., D4-GDI and Bcl-2, and in the nucleus (e.g. PARP).  This antibody has been reported to specifically recognize the active form of caspase-3 in human and mouse cells.  It has not been reported to recognize the pro-enzyme form of caspase-3.

550557 Rev. 4
Format Details
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Biotin
Biotin is a ubiquitous co-factor (also known as Vitamin B7) that has many properties that make it extremely useful for molecular biology. Biotin has an extremely high affinity for the Avidin family of proteins (Kd = 10-15 M), making it the perfect tool to link two molecules. Biotin labeled antibodies can be combined with any number of Avidin-conjugated probes in order to customize an assay to a particular need. This is especially useful in the case of magnetic cell separation using streptavidin/magnetic bead conjugates, or in the case of flow cytometry using streptavidin/fluorophore conjugates.
Biotin
550557 Rev.4
Citations & References
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Development References (3)

  1. Dai C, Krantz SB. Interferon gamma induces upregulation and activation of caspases 1, 3, and 8 to produce apoptosis in human erythroid progenitor cells. Blood. 1999; 93(10):3309-3316. (Biology). View Reference
  2. Fujita N, Tsuruo T. Involvement of Bcl-2 cleavage in the acceleration of VP-16-induced U937 cell apoptosis. Biochem Biophys Res Commun. 1998; 246(2):484-488. (Biology). View Reference
  3. Thornberry NA, Lazebnik Y. Caspases: enemies within. Science. 1998; 281(5381):1312-1316. (Biology). View Reference
550557 Rev. 4

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.