Product Pathways - DNA Damage

Phospho-p53 Antibody Sampler Kit #9919

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Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IF-IC=Immunofluorescence (Immunocytochemistry)  IC-ABC=Immunocytochemistry (ABC)  IC=Immunocytochemistry  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  Hm=Hamster

Specificity / Sensitivity

Phospho-p53 (Ser6), (Ser9), (Ser15), (Ser20), (Ser37) and (Ser392) Antibodies detect p53 only when phosphorylated at the indicated sites and do not cross-react with p53 phosphorylated at other sites.

Western Blotting

Western blot analysis of p53 fusion protein with and without CKII phosphorylation, using Phospho-p53 (Ser392) Antibody #9281 or p53 Antibody #9282. Phospho-p53 Antibody reacts specifically with as little as 1 ng of phosphorylated p53 protein.

Western Blotting

Western blot analysis of extracts from hydroxyurea (20 mM) treated MvILu cells, using Phospho-p53 (Ser392) Antibody #9281.

Western Blotting

Western analysis of E. coli and Baculovirus expressed p53 fusion protein, using p53 Antibody #9282.

Western Blotting

Western blot analysis of extracts from 293, COS, HeLa, A431 and NBT-II cells with and without treatment, using p53 Antibody #9282.

Western Blotting

Western blot analysis of extracts from UV or hydroxyurea (20 mM) treated MvILu cells, using Phospho-p53 (Ser15) Antibody #9284.

Western Blotting

Specificity and sensitivity of Phospho-p53 (Ser15) Antibody #9284. Western blot analysis of p53 fusion protein with and without DNA-PK phosphorylation, using Phospho-p53 (Ser15) Antibody (upper) and p53 Antibody #9282 (lower).

Western Blotting

Western blot analysis of extracts from UV treated PC12 cells or HeLa cells, using Phospho-p53 (Ser15) Antibody #9284.

Western Blotting

Western blot analysis of extracts from COS cells treated with UV or MMS, using Phospho-p53 (Ser6) Antibody #9285 or p53 Antibody #9282.

Western Blotting

Western blot analysis of p53 fusion protein with and without CKI or CKII phosphorylation, using Phospho-p53 (Ser6) Antibody #9285 or p53 Antibody #9282.

Western Blotting

Western blot analysis of cell extracts from hydroxyurea (20 mM) or UV treated MvlLu cells, using Phospho-p53 (Ser15) 16G8 Monoclonal Antibody #9286.

Western Blotting

Western blot analysis of extracts from COS cells treated with UV or MMS and 293 cells treated with UV, using Phospho-p53 (Ser37) #9289 (upper) and p53 Antibody #9282 (lower).

IHC-P (paraffin)

Nuclear localization of phosphorylated p53 in paraffin-embedded human breast carcinoma, demonstrated by staining with Phospho-p53 (Ser392) Antibody #9281.

IHC-P (paraffin)

Immunohistochemical staining of paraffin-embedded human breast carcinoma, showing nuclear localization of phosphorylated p53, using Phospho-p53 (Ser15) Antibody #9284.

IHC-P (paraffin)

Immunohistochemical staining of phosphorylated p53 in paraffin-embedded human breast carcinoma showing nuclear localization, using Phospho-p53 (Ser20) Antibody #9287.

IHC-F (frozen)

Immunocytochemical staining of COS-7 cells, using p53 Antibody #9282.

IHC-F (frozen)

Immunocytochemical staining of untreated and UV treated HeLa cells, using Phospho-p53 (Ser15) Antibody #9284.

IC-ABC

Immunocytochemical staining of untreated or hydroxyurea treated MvlLu cells, using Phospho-p53 (Ser392) Antibody #9281.

IC-ABC

Immunocytochemical staining of untreated and UV treated HeLa cells, using Phospho-p53 (Ser15) Antibody #9284.

IC-ABC

Immunocytochemical staining of control or UV treated COS cells, using Phospho-p53 (Ser6) Antibody #9285.

IC-ABC

Immunocytochemical staining of untreated or hydroxyurea treated MvlLu cells (#9286).

IC-ABC

Immunocytochemical staining of control or UV treated COS cells, using Phospho-p53 (Ser20) Antibody #9287.

IC-ABC

Immunocytochemical staining of control or UV treated COS-7 cells, using Phospho-p53 (Ser37) Antibody #9289.

Source / Purification

Polyclonal antibodies are produced by immunizing rabbits with synthetic phospho-peptides (KLH coupled) corresponding to residues surrounding Ser6, Ser9, Ser15, Ser20, Ser37 and Ser392 of human p53. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibody (isotype: IgG1) is produced by immunizing mice with a synthetic phospho-Ser15 peptide (KLH coupled) corresponding to residues surrounding Ser15 of human p53. Antibody is supplied in HEPES buffer with 50% glycerol and less than 0.02% sodium azide.

Background

The p53 tumor suppressor protein plays a major role in cellular response to DNA damage and other genomic aberrations. Activation of p53 can lead to either cell cycle arrest and DNA repair or apoptosis (1). p53 is phosphorylated at multiple sites in vivo and by several different protein kinases in vitro (2,3). DNA damage induces phosphorylation of p53 at Ser15 and Ser20 and leads to a reduced interaction between p53 and its negative regulator, oncoprotein MDM2 (4). MDM2 inhibits p53 accumulation by targeting it for ubiquitination and proteasomal degradation (6,7). p53 can apparently be phosphorylated by ATM, ATR and DNA-PK at Ser15 and Ser37. Phosphorylation impairs the ability of MDM2 to bind p53, promoting both the accumulation and activation of p53 in response to DNA damage (4,5). Chk2 and Chk1 can phosphorylate p53 at Ser20, enhancing its tetramerization, stability and activity (8,9). p53 is phosphorylated at Ser392 in vivo (11,12) and by CAK in vitro (12). Phosphorylation of p53 at Ser392 is altered in human tumors (14) and has been reported to influence the growth suppressor function, DNA binding and transcriptional activation of p53 (10,11,13). p53 is phosphorylated at Ser6 and Ser9 by CK1δ and CK1ε both in vitro and in vivo (10,15). Phosphorylation of p53 at Ser46 regulates the ability of p53 to induce apoptosis (16). Acetylation of p53 is mediated by p300 and CBP acetyltransferases. Inhibition of deacetylation suppressing MDM2 from recruiting HDAC1 complex by p19 (ARF) stabilizes p53. Acetylation appears to play a positive role in the accumulation of p53 protein in stress response (17). Following DNA damage, human p53 becomes acetylated at Lys382 (Lys379 in mouse) in vivo to enhance p53-DNA binding (18). Deacetylation of p53 occurs through interaction with the SIRT1 protein, a deacetylase that may be involved in cellular aging and the DNA damage response (19).

1. Levine, A.J. (1997) Cell 88, 323-331.
2. Meek, D.W. (1994) Semin. Cancer Biol. 5, 203-210.
3. Milczarek, G.J. et al. (1997) Life Sci. 60, 1-11.
4. Shieh, S.Y. et al. (1997) Cell 91, 325-334.
5. Tibbetts, R.S. et al. (1999) Genes Dev. 13, 152-157.
6. Chehab, N.H. et al. (1999) Proc. Natl. Acad. Sci. USA 96, 13777-13782.
7. Honda, R. et al. (1997) FEBS Lett. 420, 25-27.
8. Shieh, S.Y. et al. (1999) EMBO J. 18, 1815-1823.
9. Hirao, A. et al. (2000) Science 287, 1824-1827.
10. Kohn, K.W. (1999) Mol. Biol. Cell 10, 2703-2734.
11. Hao, M. et al. (1996) J. Biol. Chem. 271, 29380-29385.
12. Lu, H. et al. (1997) Mol. Cell. Biol. 17, 5923-5934.
13. Lohrum, M. and Scheidtmann, K.H. (1996) Oncogene 13, 2527-2539.
14. Ulrich, S.J. et al. (1993) Proc. Natl. Acad. Sci. USA 90, 5954-5958.
15. Knippschild, U. et al. (1997) Oncogene 15, 1727-1736.
16. Oda, K. et al. (2000) Cell 102, 849-862.
17. Ito, A. et al. (2001) EMBO J. 20, 1331-1340.
18. Sakaguchi, K. et al. (1998) Genes Dev. 12, 2831-2841.
19. Solomon, J.M. et al. (2006) Mol. Cell. Biol. 26, 28-38.

Application References

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