PAR/pADPr Antibody

  
  • Specificity
    This polyclonal antibody detects free PAR and poly-ribosylated proteins.
  • Source
    Polyclonal Rabbit IgG
  • Purification
    Antigen Affinity-purified
  • Immunogen
    Poly(ADP-ribose) polymer
  • Formulation
    This antibody is an affinity purified IgG fraction in 1X PBS, containing 50% glycerol.
  • Label
    Unconjugated
Applications
  •  
    Recommended
    Concentration
    Sample
  • Western Blot
    1:1000 dilution
    See below
Please Note: Optimal dilutions should be determined by each laboratory for each application. General Protocols are available in the Technical Information section on our website.
Data Examples
Detection of PAR/pADPr by Western Blot. Western blot analysis of Wehi cellsuntreated (H) and treated (T) with 25 μM Etoposidefor 4 hours at 37 °C. Cells were lysed in Tris-GlycineSDS sample buffer at the concentration of 1 x 107cells/ml, and 10 μl of clarified lysate were loadedper well of 4-20% Tris-Glycine gel. Proteins weretransferred onto an Immobilon FL membrane andribosylated proteins were detected using Trevigen'spolyclonal anti-PAR antibody (cat# 4336-APC-050)followed by an IR680-conjugated secondary antibody(Licor). The membrane was scanned using anOdyssey Infrared Imaging System (Licor).
Preparation and Storage
  • Shipping
    The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
  • Stability & Storage
    -20°C (manual defrost freezer)
Background: PAR/pADPr
Procedure for Immunoblotting using Peroxidase Detection:

Blotting buffer: 12 mM Tris base, 96 mM Glycine, and 15% MeOH.
Blocking solution: 5% (w/v) nonfat dry milk in TBS-0.1% Tween.
Antibody solution: 5% (w/v) nonfat dry milk, in TBS-0.1% Tween.

Transfer the electrophoresed proteins to a PVDF membrane and incubate the membrane for 1 hour at room temperature in blocking solution.

Incubate the membrane overnight at 4°C in antibody solution containing a 1:1000 dilution of anti-PAR rabbit polyclonal antibody. Empirical determination of primary antibody concentration will be required for optimal results.

Wash the membrane at room temperature for 5 minutes with 3 changes of TBS-0.1% Tween. Changing the membrane containers often reduces background.

Incubate the membrane at room temperature for 1 hour in antibody solution containing antirabbit conjugated to horseradish peroxidase.

Empirical determination of secondary antibody concentration will be required for optimal results.

Wash the membrane for 5 minutes with 4 changes of TBS-0.1% Tween.

Develop peroxidase reaction using chemiluminescence.
  • References:
    1. Affar, E.B., et al. 1998. Immunodot blot method for the detection of poly(ADP-ribose) synthesized in vitro and in vivo. Anal Biochem 259:280-3.
    2. Shah, G.M., et al. 1995. Methods for biochemical study of poly(ADP-ribose) metabolism in vitro and in vivo. Anal Biochem 227:1-13.
    3. Gagné, J-P., et al. 2008. Proteome-wide identification of poly(ADP-ribose) binding proteins and poly(ADP-ribose)-associated protein complexes. Nucleic Acids Res 36:6959-76.
  • Long Name:
    Poly [ADP-ribose] Polymer
  • Alternate Names:
    pADPr; PAR
Related Research Areas
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