Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

Acetyl-Histone H3 (Lys9) Antibody #9671

Applications Reactivity Sensitivity MW (kDa) Source
W IP IHC-P ChIP H M R Mk Dm Sc Endogenous 17 Rabbit

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  Dm=D. melanogaster  Sc=S. cerevisiae
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Acetyl-Histone H3 (Lys9) Antibody detects endogenous levels of Histone H3 only when acetylated at lysine 9. It does not cross-react with phosphorylated histone H3.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic acetylated peptide corresponding to residues surrounding Lys9 of histone H3. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from NIH/3T3 cells, untreated, TSA-treated (to induce acetylation), serum and calyculin A-treated (to induce phosphorylation), or both, using Acetyl-Histone H3 (Lys9) Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, showing nuclear localization, using Acetyl-Histone H3 (Lys9) Antibody.

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma, using Acetyl-Histone H3 (Lys 9) Antibody.


IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human colon carcinoma, using Acetyl-Histone H3 (Lys9) Antibody in the presence of control peptide (left) or Acetyl-Histone H3 (Lys9) blocking peptide #1083 (right).

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 10 μl of Acetyl-Histone H3 (Lys9) Antibody or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human RPL30 Exon 3 Primers #7014, SimpleChIP® Human AFM Intron 1 Primers #5098, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.

Background

Modulation of chromatin structure plays an important role in the regulation of transcription in eukaryotes. The nucleosome, made up of DNA wound around eight core histone proteins (two each of H2A, H2B, H3, and H4), is the primary building block of chromatin (1). The amino-terminal tails of core histones undergo various post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (2-5). These modifications occur in response to various stimuli and have a direct effect on the accessibility of chromatin to transcription factors and, therefore, gene expression (6). In most species, histone H2B is primarily acetylated at Lys5, 12, 15, and 20 (4,7). Histone H3 is primarily acetylated at Lys9, 14, 18, 23, 27, and 56. Acetylation of H3 at Lys9 appears to have a dominant role in histone deposition and chromatin assembly in some organisms (2,3). Phosphorylation at Ser10, Ser28, and Thr11 of histone H3 is tightly correlated with chromosome condensation during both mitosis and meiosis (8-10). Phosphorylation at Thr3 of histone H3 is highly conserved among many species and is catalyzed by the kinase haspin. Immunostaining with phospho-specific antibodies in mammalian cells reveals mitotic phosphorylation at Thr3 of H3 in prophase and its dephosphorylation during anaphase (11).

  1. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  2. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  3. Strahl, B.D. and Allis, C.D. (2000) Nature 403, 41-5.
  4. Cheung, P. et al. (2000) Cell 103, 263-71.
  5. Bernstein, B.E. and Schreiber, S.L. (2002) Chem Biol 9, 1167-73.
  6. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  7. Thorne, A.W. et al. (1990) Eur J Biochem 193, 701-13.
  8. Hendzel, M.J. et al. (1997) Chromosoma 106, 348-60.
  9. Goto, H. et al. (1999) J Biol Chem 274, 25543-9.
  10. Preuss, U. et al. (2003) Nucleic Acids Res 31, 878-85.
  11. Dai, J. et al. (2005) Genes Dev 19, 472-88.

Application References

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For Research Use Only. Not For Use In Diagnostic Procedures.

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