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    BV421 Mouse Anti-Cleaved PARP (Asp 214)
    BV421 Mouse Anti-Cleaved PARP (Asp 214)
    Flow cytometric analysis of cleaved PARP (Asp 214) expression (left panel). Cells from the human Jurkat (ATCC TIB-152) cell line were cultured for 6 hours with (solid line histogram) or without (dashed line histogram) Camptothecin (Sigma-Aldrich Cat. No. C-9911; 12 μM final concentration). The cells were harvested, washed with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656), and fixed and permeabilized with 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 BD Horizon™ BV421 Mouse Anti-Cleaved PARP (Asp 214) antibody (Cat. No. 564129). Histograms were derived from gated events with the forward and side light-scatter characteristics of intact Jurkat cells. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System. Immunofluorescent analysis of cleaved PARP in apoptotic human cells (right panel). HeLa cells (ATCC, CCL-2) were treated with camptothecin (20 µM, 6 hours) to induce apoptosis. Cells were then fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050), and blocked with 5% goat serum, 1% BSA, and 0.5% Triton™ X-100 diluted in PBS. Cells were stained with BD Horizon™ BV421 Mouse Anti-Cleaved PARP (Asp 214) antibody (Cat. No. 564129, pseudo-colored red) and Alexa Fluor® 488 Mouse Anti-GM130 antibody (Cat. No. 560257, pseudo-colored green). DRAQ5 was used as a nuclear counterstain (Cat. No. 564902/564903, pseudo-colored blue). Images were captured on a standard epifluorescence microscope. Original magnification, 20x.
    BV421 Mouse Anti-Cleaved PARP (Asp 214)
    Flow cytometric analysis of cleaved PARP (Asp 214) expression (left panel). Cells from the human Jurkat (ATCC TIB-152) cell line were cultured for 6 hours with (solid line histogram) or without (dashed line histogram) Camptothecin (Sigma-Aldrich Cat. No. C-9911; 12 μM final concentration). The cells were harvested, washed with BD Pharmingen™ Stain Buffer (FBS) (Cat. No. 554656), and fixed and permeabilized with 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 BD Horizon™ BV421 Mouse Anti-Cleaved PARP (Asp 214) antibody (Cat. No. 564129). Histograms were derived from gated events with the forward and side light-scatter characteristics of intact Jurkat cells. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System. Immunofluorescent analysis of cleaved PARP in apoptotic human cells (right panel). HeLa cells (ATCC, CCL-2) were treated with camptothecin (20 µM, 6 hours) to induce apoptosis. Cells were then fixed with BD Cytofix™ Fixation Buffer (Cat. No. 554655), permeabilized with BD Phosflow™ Perm Buffer III (Cat. No. 558050), and blocked with 5% goat serum, 1% BSA, and 0.5% Triton™ X-100 diluted in PBS. Cells were stained with BD Horizon™ BV421 Mouse Anti-Cleaved PARP (Asp 214) antibody (Cat. No. 564129, pseudo-colored red) and Alexa Fluor® 488 Mouse Anti-GM130 antibody (Cat. No. 560257, pseudo-colored green). DRAQ5 was used as a nuclear counterstain (Cat. No. 564902/564903, pseudo-colored blue). Images were captured on a standard epifluorescence microscope. Original magnification, 20x.
    Product Details
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    BD Horizon™
    Asp214
    Human (QC Testing), Mouse (Tested in Development)
    Mouse IgG1, κ
    Human cleaved PARP
    Intracellular staining (flow cytometry) (Routinely Tested), Immunocytochemistry, Immunofluorescence (Tested During Development)
    5 µl
    142, 11545
    AB_2738611
    Aqueous buffered solution containing BSA 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 BV421 under optimum conditions, and unconjugated antibody and free BD Horizon BV421 were removed.
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    Recommended Assay Procedures

    For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant™ dyes are used in a multicolor flow cytometry panel.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

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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. Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
    6. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
    7. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
    8. BD Horizon Brilliant Violet 421 is covered by one or more of the following US patents: 8,158,444; 8,362,193; 8,575,303; 8,354,239.
    9. BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
    10. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
    11. Triton is a trademark of the Dow Chemical Company.
    12. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
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    564129 Rev. 2
    Antibody Details
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    F21-852

    PARP (Poly [ADP-Ribose] Polymerase) is a 113-kDa nuclear chromatin-associated enzyme that catalyzes the transfer of ADP-ribose units from NAD+ to a variety of nuclear proteins including topoisomerases, histones, and PARP itself.  The catalytic activity of PARP is increased in cells following DNA damage, and PARP is thought to play an important role in mediating the normal cellular response to DNA damage.  Additionally, PARP is a target of the caspase protease activity associated with apoptosis.  The PARP protein consists of an N-terminal DNA-binding domain (DBD) and a C-terminal catalytic domain separated by a central automodification domain.  During apoptosis, Caspase-3 cleaves PARP at a recognition site (Asp Glu Val Asp Gly) in the DBD to form 24- and 89-kDa fragments.  This process separates the DBD (which is mostly in the 24-kDa fragment) from the catalytic domain (in the 89-kDa fragment) of the enzyme, resulting in the loss of normal PARP function.  It has been proposed that inactivation of PARP directs DNA-damaged cells to undergo apoptosis rather than necrotic degradation, and the presence of the 89-kDa PARP cleavage fraction is considered to be a marker of apoptosis.

    A peptide corresponding to the N-terminus of the cleavage site (Asp 214) of human PARP was used as the immunogen. The F21-852 monoclonal antibody reacts only with the 89-kDa fragment of human PARP-1 that is downstream of the Caspase-3 cleavage site (Asp214) and contains the automodification and catalytic domains.  It does not react with intact human PARP-1.  Cross-reactivity with other members of the PARP superfamily is unknown.  Recognition of cleaved PARP in mouse cells has been demonstrated, and it may also cross-react with a number of other species due to the conserved nature of the molecule.

    564129 Rev. 2
    Format Details
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    BV421
    The BD Horizon Brilliant Violet™ 421 (BV421) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This polymer-technology based dye has an excitation maximum (Ex Max) of 407-nm and an emission maximum (Em Max) at 423-nm. Driven by BD innovation, BV421 is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 420-nm (e.g., a 431/28-nm or 450/50-nm bandpass filter). BV421 is an ideal alternative for V450 as it is approximately ten times brighter with less spillover into the BV510/V500 detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
    altImg
    BV421
    Violet 405 nm
    407 nm
    423 nm
    564129 Rev.2
    Citations & References
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    Development References (10)

    1. Amé J-C, Spenlehauer C, de Murcia G. The PARP superfamily. Bioessays. 2004; 26:882-893. (Biology).
    2. Boulares AH, Yakovlev AG, Ivanova V, et al. Role of Poly(ADP-ribose) polymerase (PARP) cleavage in apoptosis. Caspase 3-resistant PARP mutant increases rates of apoptosis in transfected cells. J Biol Chem. 1999; 274(33):22932-22940. (Biology).
    3. Cherney BW, McBride OW, Chen D, et al. cDNA sequence, protein structure, and chromosomal location of the human gene for poly(ADP-ribose) polymerase. Proc Natl Acad Sci U S A. 1987; 84(23):8370-8374. (Biology). View Reference
    4. D'Amours D, Desnoyers S, D'Silva I, Poirier GG. Poly(ADP-ribosyl)ation reactions in the regulation of nucelar functions. Biochem J. 1999; 342:249-268. (Biology).
    5. Kaufmann SH, Desnoyers S, Ottaviano Y, Davidson NE, Poirier GG. Specific proteolytic cleavage of poly(ADP-ribose) polymerase: an early marker of chemotherapy-induced apoptosis. Cancer Res. 1993; 53(17):3976-3985. (Biology). View Reference
    6. Lamarre D, Talbot B, Leduc Y, Muller S, Poirier G. Production and characterization of monoclonal antibodies specific for the functional domains of poly(ADP-ribose) polymerase. Biochem Cell Biol. 1986; 64(4):368-376. (Biology). View Reference
    7. Lamarre D, Talbot B, de Murcia G, et al. Structural and functional analysis of poly(ADP ribose) polymerase: an immunological study. Biochim Biophys Acta. 1988; 950(2):147-160. (Biology). View Reference
    8. Patel T, Gores GJ, Kaufmann SH. The role of proteases during apoptosis. FASEB J. 1996; 10(5):587-597. (Biology). View Reference
    9. Soldani G, Scovassi AI. Poly(ADP-ribose) polymerase-1 cleavage during apoptosis: an update. Apoptosis. 2002; 7:321-328. (Biology).
    10. Tewari M, Quan LT, O'Rourke K, et al. Yama/CPP32 beta, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly(ADP-ribose) polymerase. Cell. 1995; 81(5):801-809. (Biology). View Reference
    View All (10) View Less
    564129 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.

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