Revision 4
Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

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

WB, IP, FC-FP, ChIP, ChIP-seq

REACTIVITY:

H M R

SENSITIVITY:

Endogenous

MW (kDa):

14

Source/Isotype:

Rabbit IgG

UniProt ID:

#P0C0S5

Entrez-Gene Id:

3015

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
Immunoprecipitation 1:200
Flow Cytometry (Fixed/Permeabilized) 1:50
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 H2AZ (Lys4/Lys7) recognizes endogenous levels of histone H2AZ protein only when acetylated at Lys4 and/or Lys7. This antibody does not cross-react with other acetylated histones, including histone H2A acetylated at Lys5. This antibody also detects a band around 22 kDa, which is most likely monoubiquitylated histone H2AZ that is acetylated on Lys4 and Lys7.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding acetylated Lys4 and Lys7 of human H2AZ protein.

Background

Modulation of chromatin structure plays a critical role in the regulation of transcription in eukaryotes. The nucleosome, made up of four core histone proteins (H2A, H2B, H3 and H4), is the primary building block of chromatin. In addition to the growing number of post-translational histone modifications regulating chromatin structure, cells can also exchange canonical histones with variant histones that can directly or indirectly modulate chromatin structure (1). There are five major variants of histone H2A: canonical H2A (most abundant), H2A.X, MacroH2A, H2ABbd and H2A.Z (2). Histone H2A.Z, the most conserved variant across species, functions as both a positive and negative regulator of transcription and is important for chromosome stability (2). Several homologous protein complexes, such as SWR-C (S. cerevisiae), TIP60 (D. melanogaster) and SRCAP (mammals), have been shown to catalyze the ATP-dependent exchange of H2A.Z for H2A in the nucleosome (3,4,5). This exchange of histone H2A variants changes histone-histone interactions in the nucleosome core and alters an acidic patch on the surface of the nucleosome, resulting in changes in nucleosome stability and binding of non-histone proteins such as HP1α (6,7).

Acetylation of Histone H2AZ correlates with gene activity (8). Acetylation of Histone H2AZ on Lys4 and Lys7 occurs at the 5' end of genes and confers nucleome destabilization and open chromatin confirmation required for tanscriptional activation (9-11).

  1. Jin, J. et al. (2005) Trends Biochem Sci 30, 680-7.
  2. Raisner, R.M. and Madhani, H.D. (2006) Curr Opin Genet Dev 16, 119-24.
  3. Mizuguchi, G. et al. (2004) Science 303, 343-8.
  4. Kusch, T. et al. (2004) Science 306, 2084-7.
  5. Ruhl, D.D. et al. (2006) Biochemistry 45, 5671-7.
  6. Suto, R.K. et al. (2000) Nat Struct Biol 7, 1121-4.
  7. Fan, J.Y. et al. (2004) Mol Cell 16, 655-61.
  8. Millar, C.B. et al. (2006) Genes Dev 20, 711-22.
  9. Bruce, K. et al. (2005) Nucleic Acids Res 33, 5633-9.
  10. Ishibashi, T. et al. (2009) Biochemistry 48, 5007-17.
  11. Valdés-Mora, F. et al. (2012) Genome Res 22, 307-21.

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 IP: Immunoprecipitation 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.
SimpleChIP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

使用に関する制限

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Revision 4
#75336

Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb

Western Blotting Image 1: Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb Expand Image
Western blot analysis of extracts from HeLa cells, untreated (-) or treated (+) with Trichostatin A (TSA) #9950 (1 μM, 18 hr), using Acetyl-Histone H2A.Z (Lys4/Lys7) Rabbit mAb (upper) or Histone H2A.Z Antibody #2718 (lower).
No image available
Flow Cytometry Image 1: Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb Expand Image
Flow cytometric analysis of HeLa cells, untreated (blue) or treated with Trichostatin A (TSA) #9950 (1 µM, 18 hr; green) using Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Chromatin Immunoprecipitation Image 1: Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells treated with Trichostatin A (TSA) #9950 (100 nM, 24 hr) and Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Library was prepared using DNA Library Prep Kit for Illumina (ChIP-seq, CUT&RUN) #56795. The figure shows binding across GAPDH gene.
Chromatin Immunoprecipitation Image 2: Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells treated with Trichostatin A (TSA) #9950 (100 nM, 24 hr) and Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Library was prepared using DNA Library Prep Kit for Illumina (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 12 (upper), including GAPDH gene (lower).
Chromatin Immunoprecipitation Image 3: Acetyl-Histone H2A.Z (Lys4/Lys7) (D3V1I) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from LNCaP cells treated with Trichostatin A (TSA) #9950 (100 nM, 24 hr) and either Acetyl-Histone H2A.Z (Lys4/7) (D3V1I) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by Real-Time PCR using human CAV1 promoter primers, SimpleChIP® Human KLK2 Intron 1 Primers #62086, and SimpleChIP® Human α Satelite 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.