Revision 6
Cell Signaling Technology

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3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, W-S, IP, IF-IC, FC-FP, ChIP, ChIP-seq

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

17

Source/Isotype:

Rabbit IgG

UniProt ID:

#P68431

Entrez-Gene Id:

8350

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.
Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10 - 1:50
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:12800 - 1:25600
Flow Cytometry (Fixed/Permeabilized) 1:1600 - 1:6400
Chromatin IP 1:50
Chromatin IP-seq 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Specificity / Sensitivity

Acetyl-Histone H3 (Lys14) (D4B9) Rabbit mAb recognizes endogenous levels of Histone H3 protein only when acetylated at Lys14. This antibody does not cross react with histone H3 acetylated at Lys9, 18, 27, or 56.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Hamster, D. melanogaster, Xenopus, Zebrafish, Pig, S. cerevisiae, Horse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding acetylated Lys14 of human Histone H3 protein.

Background

The nucleosome, made up of four core histone proteins (H2A, H2B, H3, and H4), is the primary building block of chromatin. Originally thought to function as a static scaffold for DNA packaging, histones have now been shown to be dynamic proteins, undergoing multiple types of post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (1,2). Histone acetylation occurs mainly on the amino-terminal tail domains of histones H2A (Lys5), H2B (Lys5, 12, 15, and 20), H3 (Lys9, 14, 18, 23, 27, 36, and 56), and H4 (Lys5, 8, 12, and 16) and is important for the regulation of histone deposition, transcriptional activation, DNA replication, recombination, and DNA repair (1-3). Hyper-acetylation of the histone tails neutralizes the positive charge of these domains and is believed to weaken histone-DNA and nucleosome-nucleosome interactions, thereby destabilizing chromatin structure and increasing the accessibility of DNA to various DNA-binding proteins (4,5). In addition, acetylation of specific lysine residues creates docking sites for a protein module called the bromodomain, which binds to acetylated lysine residues (6). Many transcription and chromatin regulatory proteins contain bromodomains and may be recruited to gene promoters, in part, through binding of acetylated histone tails. Histone acetylation is mediated by histone acetyltransferases (HATs), such as CBP/p300, GCN5L2, PCAF, and Tip60, which are recruited to genes by DNA-bound protein factors to facilitate transcriptional activation (3). Deacetylation, which is mediated by histone deacetylases (HDAC and sirtuin proteins), reverses the effects of acetylation and generally facilitates transcriptional repression (7,8).

  1. Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
  2. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  3. Roth, S.Y. et al. (2001) Annu Rev Biochem 70, 81-120.
  4. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  5. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  6. Yang, X.J. (2004) Bioessays 26, 1076-87.
  7. Haberland, M. et al. (2009) Nat Rev Genet 10, 32-42.
  8. Haigis, M.C. and Sinclair, D.A. (2010) Annu Rev Pathol 5, 253-95.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting W-S: Simple Western™ IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
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