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V450 Rabbit Anti-Active Caspase-3
V450 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 (shaded) or were treated with 4-12 µM of camptothecin (Sigma-Aldrich Cat. No. C9911) for 4-6 hr to induce apoptosis (unshaded). Cells were washed once in 1X PBS, then fixed and permeabilized using the

BD Cytofix/Cytoperm™ Kit (Cat. No. 554714) followed by staining with the BD Horizon™ V450 Rabbit Anti-Active Caspase-3 antibody.  Histograms were derived from gated events based on light scattering characteristics for Jurkat cells.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.

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 (shaded) or were treated with 4-12 µM of camptothecin (Sigma-Aldrich Cat. No. C9911) for 4-6 hr to induce apoptosis (unshaded). Cells were washed once in 1X PBS, then fixed and permeabilized using the

BD Cytofix/Cytoperm™ Kit (Cat. No. 554714) followed by staining with the BD Horizon™ V450 Rabbit Anti-Active Caspase-3 antibody.  Histograms were derived from gated events based on light scattering characteristics for Jurkat cells.  Flow cytometry was performed on a BD™ LSR II flow cytometry system.

Product Details
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BD Horizon™
CPP32; Yama; Apopain
Human (QC Testing), Mouse (Predicted)
Rabbit IgG
Human Active Caspase-3 Fragment
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_1727415
Aqueous buffered solution containing protein stabilizer and ≤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 with BD Horizon™ V450 under optimum conditions, and unreacted BD Horizon™ V450 was removed.

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. BD Horizon V450 has a maximum absorption of 406 nm and maximum emission of 450 nm. Before staining with this reagent, please confirm that your flow cytometer is capable of exciting the fluorochrome and discriminating the resulting fluorescence.
  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. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  6. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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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.

The antibody is conjugated to BD Horizon™ V450, which has been developed for use in multicolor flow cytometry experiments and is available exclusively from BD Biosciences. It is excited by the Violet laser Ex max of 406 nm and has an Em Max at 450 nm. Conjugates with BD Horizon™ V450 can be used in place of Pacific Blue™ conjugates.

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Format Details
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V450
BD Horizon™ V450 Dye is part of the BD Horizon™ violet family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 405-nm and an emission maximum (Em Max) at 450-nm. BD Horizon™ V450, driven by BD innovation, is designed to be excited by the violet laser (405 nm) and detected using an optical filter centered near 450-nm (e.g., a 450/50-nm bandpass filter). The dye can be excited by the UV (355-nm) laser resulting in cross-laser excitation and spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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V450
Violet 405 nm
405 nm
450 nm
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Citations & References
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Development References (15)

  1. Alnemri ES, Livingston DJ, Nicholson DW, et al. Human ICE/CED-3 protease nomenclature. Cell. 1996; 87(2):171. (Biology). View Reference
  2. 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
  3. Donoghue S, Baden HS, Lauder I, Sobolewski S, Pringle JH. Immunohistochemical localization of caspase-3 correlates with clinical outcome in B-cell diffuse large-cell lymphoma. Cancer Res. 1999; 59(20):5386-5391. (Biology). View Reference
  4. Fernandes-Alnemri T, Litwack G, Alnemri ES. CPP32, a novel human apoptotic protein with homology to Caenorhabditis elegans cell death protein Ced-3 and mammalian interleukin-1 beta-converting enzyme. J Biol Chem. 1994; 269(49):30761-30764. (Biology). View Reference
  5. 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
  6. Harvey NL, Butt AJ, Kumar S. Functional activation of Nedd2/ICH-1 (caspase-2) is an early process in apoptosis. J Biol Chem. 1997; 272(20):13134-13139. (Biology). View Reference
  7. Jänicke RU, Sprengart ML, Wati MR, Porter AG. Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis. J Biol Chem. 1998; 273(16):9357-9360. (Biology). View Reference
  8. Li J, Chen P, Sinogeeva N, et al. Arsenic trioxide promotes histone H3 phosphoacetylation at the chromatin of CASPASE-10 in acute promyelocytic leukemia cells. J Biol Chem. 2002; 277(51):49504-49510. (Biology). View Reference
  9. Martin SJ, Finucane DM, Amarante-Mendes GP, O'Brien GA, Green DR. Phosphatidylserine externalization during CD95-induced apoptosis of cells and cytoplasts requires ICE/CED-3 protease activity. J Biol Chem. 1996; 271(46):28753-28756. (Biology). View Reference
  10. Miossec C, Dutilleul V, Fassy F, Diu-Hercend A. Evidence for CPP32 activation in the absence of apoptosis during T lymphocyte stimulation. J Biol Chem. 1997; 272(21):13459-13462. (Biology). View Reference
  11. Patel T, Gores GJ, Kaufmann SH. The role of proteases during apoptosis. FASEB J. 1996; 10(5):587-597. (Biology). View Reference
  12. Scaffidi C, Fulda S, Srinivasan A, et al. Two CD95 (APO-1/Fas) signaling pathways.. EMBO J. 1998; 17(6):1675-1687. (Biology).
  13. Suzuki Y, Nakabayashi Y, Takahashi R. Ubiquitin-protein ligase activity of X-linked inhibitor of apoptosis protein promotes proteasomal degradation of caspase-3 and enhances its anti-apoptotic effect in Fas-induced cell death. Proc Natl Acad Sci U S A. 2001; 98(15):8662-8667. (Biology). View Reference
  14. Takemoto K, Nagai T, Miyawaki A, Miura M. Spatio-temporal activation of caspase revealed by indicator that is insensitive to environmental effects. J Cell Biol. 2003; 160(2):235-243. (Biology). View Reference
  15. Thornberry NA, Lazebnik Y. Caspases: enemies within. Science. 1998; 281(5381):1312-1316. (Biology). View Reference
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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.