BD Pharmingen™ Purified Mouse Anti-Poly ADP

BD Pharmingen™
Technical Data Sheet
Purified Mouse Anti-Poly ADP-Ribose
Product Information
Material Number:
Alternate Name:
Size:
Concentration:
Clone:
Immunogen:
Isotype:
Reactivity:
Storage Buffer:
550781
PAR
50 µg
0.5 mg/ml
10H
Human Poly ADP-Ribose
Mouse IgG3, κ
QC Testing: Human
Reported: Mouse, Rat, Drosophilia
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
Description
The ability of an organism to repair DNA damage caused by oxidative stress and UV exposure correlates with the organisms' ability to survive.
One mechanism whereby eukaryotic cells respond to DNA damage is the synthesis of poly ADP-ribose (PAR). PAR is synthesized after
activation of the DNA repair enzyme poly(ADP-ribose) polymerase (PARP). Upon DNA damage, PARP binds to DNA strand breaks and
catalyses the addition of long branched chains of PAR to a number of nuclear proteins, including topoisomerases, histones, and PARP. The
amount of PAR formed in living cells with DNA damage is commensurate with the extent of the damage. Under DNA damage conditions,
PAR-fluorescence is detectable within the first minutes and then later disappears. PAR can also be detected in cells undergoing apoptosis, and
appears to be quite stable. The appearance of PAR correlates with the specific cleavage of PARP by caspase-3 and is correlative with cells
undergoing apoptosis. The antibody recognizes PAR in numerous species including, human, mouse, rat, and drosophila. Purified poly
(ADP-ribose) was used as immunogen.
Immunofluorescence microscopy. HeLa cells growing on Labtekll
chambered coverglass were either treated with 1.0 mM H2O2 at 37°C
for 10 minutes (top left) or left untreated (bottom left), washed 1x in
PBS, and fixed MEOH/acetone (70:30 v/v) for 10 minutes at -20°C.
Following fixation, cells were air dried and PBS was added to the
slide and incubated for 10 minutes at room temperature. PBS was
removed and cells were permeabilized in blocking buffer [PBS, 5%,
non-fat dry milk (w/v), 0.05% Tween 20] for 30 minutes at room
temperature. Blocking buffer was removed and cells were then
stained with purified anti-poly ADP-ribose (clone 10H, Cat. No.
550781: 5 µg/ml in blocking buffer) for 1 hour at 37°C. Cells were
washed 5x in PBS and incubated with FITC-conjugated goat
anti-mouse secondary antibody (Cat. No. 554001; 5 µg/ml/blocking
buffer) for 1 hour at room temperature. Cells were washed 5x in PBS
and visualized by immunofluorescence microscopy. Top right and
bottom right panels represent phase contrast correlates of top left and
bottom left panels, respectively.
Preparation and Storage
The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
Store undiluted at 4°C.
550781 Rev. 2
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Application Notes
Application
Immunofluorescence
Routinely Tested
Western blot
Reported
Recommended Assay Procedure:
Applications include immunofluorescence microscopy (5 µg/ml) and western blot. The antibody is routinely tested by immunofluorescence
microscopy on HeLa cells treated with the DNA damaging agent H2O2. Western Blot application is reported.
Suggested Companion Products
Catalog Number
554001
Name
FITC Goat Anti-Mouse Ig
Size
0.5 mg
Clone
Polyclonal
Product Notices
1.
2.
3.
4.
5.
Since applications vary, each investigator should titrate the reagent to obtain optimal results.
Please refer to www.bdbiosciences.com/pharmingen/protocols for technical protocols.
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.
Source of all serum proteins is from USDA inspected abattoirs located in the United States.
Sodium azide is a reversible inhibitor of oxidative metabolism; therefore, antibody preparations containing this preservative agent must not
be used in cell cultures nor injected into animals. Sodium azide may be removed by washing stained cells or plate-bound antibody or
dialyzing soluble antibody in sodium azide-free buffer. Since endotoxin may also affect the results of functional studies, we recommend the
NA/LE (No Azide/Low Endotoxin) antibody format, if available, for in vitro and in vivo use.
References
Alvarez-Gonzalez R, Spring H, Müller M, Bürkle A. Selective loss of poly(ADP-ribose) and the 85-kDa fragment of poly(ADP-ribose) polymerase in nucleoli during
alkylation-induced apoptosis of HeLa cells. J Biol Chem. 1999; 5;274(45):32122-32126. (Clone-specific: Immunofluorescence, Western blot)
Donzelli M, Negri C, Mandarino A, et al. Poly(ADP-ribose) synthesis: a useful parameter for identifying apoptotic cells. Histochem J. 1997; 29(11-12):831-837.
(Biology)
Kawamitsu H, Hoshino H, Okada H, Miwa M, Momoi H, Sugimura T. Monoclonal antibodies to poly(adenosine diphosphate ribose) recognize different structures.
Biochemistry. 1984; 23(16):3771-3777. (Immunogen)
Kupper JH, van Gool L, Muller M, Burkle A. Detection of poly(ADP-ribose) polymerase and its reaction product poly(ADP-ribose) by immunocytochemistry.
Histochem J. 1996; 28(5):391-395. (Biology)
Negri C, Donzelli M, Bernardi R, Rossi L, Burkle A, Scovassi AI. Multiparametric staining to identify apoptotic human cells. Exp Cell Res. 1997; 234(1):174-177.
(Biology)
Okita N, Ashizawa D, Ohta R, Abe H, Tanuma S. Discovery of novel poly(ADP-ribose) glycohydrolase inhibitors by a quantitative assay system using dot-blot with
anti-poly(ADP-ribose). Biochem Biophys Res Commun. 2010; 392(4):485-489. (Clone-specific: Western blot)
Patel T, Gores GJ, Kaufmann SH. The role of proteases during apoptosis. FASEB J. 1996; 10(5):587-597. (Biology)
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