Anti-Histone H3 (acetyl K9) antibody - ChIP Grade (ab10812)

Overview

  • Product nameAnti-Histone H3 (acetyl K9) antibody - ChIP GradeSee all Histone H3 primary antibodies ...
  • Description
    Rabbit polyclonal to Histone H3 (acetyl K9) - ChIP Grade
  • SpecificitySpecific for acetyl K9, but cross-reacts slightly with K27.
  • Tested applicationsIHC-Fr, IHC-P, ChIP, IP, WB, CHIPseq, ICC/IF, Flow Cytmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Cow, Human, Saccharomyces cerevisiae, Arabidopsis thaliana, Caenorhabditis elegans, Fruit fly (Drosophila melanogaster), Zebrafish, Marmoset (common), Rice
    Predicted to work with: a wide range of other species
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, acetylated at K9.

  • Positive control
    • Calf Thymus Histone Preparation; Hela whole cell extract

Properties

Applications

Our Abpromise guarantee covers the use of ab10812 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-Fr 1/500.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ChIP Use 2-4 µg for 25 µg of chromatin.
IP Use at 20 µg/mg of lysate.
WB 1/500. Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa).Can be blocked with Human Histone H3 (acetyl K9) peptide (ab16635).
CHIPseq Use at an assay dependent concentration. PubMed: 22196736
ICC/IF Use a concentration of 1 µg/ml.
Flow Cyt Use at an assay dependent concentration.

Target

  • FunctionCore component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling.
  • Sequence similaritiesBelongs to the histone H3 family.
  • Developmental stageExpressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
  • Post-translational
    modifications
    Acetylation is generally linked to gene activation. Acetylation on Lys-10 (H3K9ac) impairs methylation at Arg-9 (H3R8me2s). Acetylation on Lys-19 (H3K18ac) and Lys-24 (H3K24ac) favors methylation at Arg-18 (H3R17me).
    Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
    Asymmetric dimethylation at Arg-18 (H3R17me2a) by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 (H3R8me2s) by PRMT5 is linked to gene repression. Asymmetric dimethylation at Arg-3 (H3R2me2a) by PRMT6 is linked to gene repression and is mutually exclusive with H3 Lys-5 methylation (H3K4me2 and H3K4me3). H3R2me2a is present at the 3' of genes regardless of their transcription state and is enriched on inactive promoters, while it is absent on active promoters.
    Methylation at Lys-5 (H3K4me), Lys-37 (H3K36me) and Lys-80 (H3K79me) are linked to gene activation. Methylation at Lys-5 (H3K4me) facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 (H3K79me) is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are linked to gene repression. Methylation at Lys-10 (H3K9me) is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 (H3S10ph) and acetylation of H3 and H4. Methylation at Lys-5 (H3K4me) and Lys-80 (H3K79me) require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are enriched in inactive X chromosome chromatin.
    Phosphorylated at Thr-4 (H3T3ph) by GSG2/haspin during prophase and dephosphorylated during anaphase. Phosphorylation at Ser-11 (H3S10ph) by AURKB is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at Ser-11 (H3S10ph) by RPS6KA4 and RPS6KA5 is important during interphase because it enables the transcription of genes following external stimulation, like mitogens, stress, growth factors or UV irradiation and result in the activation of genes, such as c-fos and c-jun. Phosphorylation at Ser-11 (H3S10ph), which is linked to gene activation, prevents methylation at Lys-10 (H3K9me) but facilitates acetylation of H3 and H4. Phosphorylation at Ser-11 (H3S10ph) by AURKB mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. Phosphorylation at Ser-11 (H3S10ph) is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylated at Ser-29 (H3S28ph) by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation. Phosphorylation at Thr-7 (H3T6ph) by PRKCBB is a specific tag for epigenetic transcriptional activation that prevents demethylation of Lys-5 (H3K4me) by LSD1/KDM1A. At centromeres, specifically phosphorylated at Thr-12 (H3T11ph) from prophase to early anaphase, by DAPK3 and PKN1. Phosphorylation at Thr-12 (H3T11ph) by PKN1 is a specific tag for epigenetic transcriptional activation that promotes demethylation of Lys-10 (H3K9me) by KDM4C/JMJD2C. Phosphorylation at Tyr-42 (H3Y41ph) by JAK2 promotes exclusion of CBX5 (HP1 alpha) from chromatin.
    Monoubiquitinated by RAG1 in lymphoid cells, monoubiquitination is required for V(D)J recombination (By similarity). Ubiquitinated by the CUL4-DDB-RBX1 complex in response to ultraviolet irradiation. This may weaken the interaction between histones and DNA and facilitate DNA accessibility to repair proteins.
  • Cellular localizationNucleus. Chromosome.
  • Information by UniProt
  • Database links
  • Alternative names
    • H 3 antibody
    • H3 3 Like Sequence MH921 antibody
    • H3 3A antibody
    • H3 3A antibody
    • H3 antibody
    • H3 Histone antibody
    • H3 Histone Family Member E Pseudogene antibody
    • H3.4 antibody
    • H3/A antibody
    • H3/g antibody
    • H31_HUMAN antibody
    • H3F3 antibody
    • H3F3 antibody
    • H3FA antibody
    • H3FT antibody
    • H3t antibody
    • HIST1H3J antibody
    • HIST3H3 antibody
    • HIST3H3 antibody
    • Histone cluster 1, H3a antibody
    • Histone H3 3 Pseudogene antibody
    • Histone H3.1 antibody
    • Histone H3.3 antibody
    • Histone H3/a antibody
    • Histone H3/b antibody
    • Histone H3/c antibody
    • Histone H3/d antibody
    • Histone H3/f antibody
    • Histone H3/h antibody
    • Histone H3/i antibody
    • Histone H3/j antibody
    • Histone H3/k antibody
    • Histone H3/l antibody
    see all

Anti-Histone H3 (acetyl K9) antibody - ChIP Grade images

  • ab10812 staining histone H3 (acetyl K9) in A549 cells treated with scriptaid (ab120883), by ICC/IF. Increase in histone H3 (acetyl K9) expression correlates with increased concentration of scriptaid, as described in literature.
    The cells were incubated at 37°C for 24 hour in media containing different concentrations of ab120883 (scriptaid) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab10812 (0.1 μg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 anti-rabbit polyclonal antibody (ab96899) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.
  • ab10812 staining Histone H3 (acetyl K9) in Mouse thyroid tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with paraformaldehyde, permeabilized with 0.1% Tween 20 and blocked with 10% serum for 30 minutes at 24°C. Samples were incubated with primary antibody (1/200 in 10% serum in PBS) for 1 hour at 24°C. An Alexa Fluor® 555-conjugated Goat anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-Histone H3 (acetyl K9) antibody - ChIP Grade (ab10812) at 1 µg/ml

    Lane 1 : Calf Thymus Histone Preparation Nuclear Lysate
    Lane 2 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (unmodified ) peptide (ab2903) at 0.5 µg/ml
    Lane 3 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (acetyl K9) peptide (ab16635) at 0.5 µg/ml
    Lane 4 : Calf Thymus Histone Preparation Nuclear Lysate with Histone H3 peptide - acetyl K14 at 0.5 µg/ml
    Lane 5 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (acetyl K18) peptide (ab24003) at 0.5 µg/ml
    Lane 6 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (acetyl K27) peptide (ab24404) at 0.5 µg/ml
    Lane 7 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (acetyl K23) peptide (ab48359) at 0.5 µg/ml

    Lysates/proteins at 0.5 µg per lane.

    Secondary
    Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/5000 dilution
    developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 15 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds
  • Chromatin was prepared from Hela cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The  ChIP was performed with 25µg of chromatin, 2µg of  ab10812 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.
        

  • ICC/IF image of ab10812 stained HeLa cells. The cells were 100% Methanol fixed (5 min) and then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab10812, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 Goat anti-Rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 100% Methanol fixed (5 min) Hek293, HepG2, and MCF-7 cells at 5µg/ml.
  • IHC image of Histone H3 (acetyl K9) staining in human breast carcinoma FFPE section, performed on a BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab10812, 1µg/ml, for 8 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
  • SK-N-SH cells fixed with 4% paraformaldehyde, permeabilized in 0.5% Triton X-100 and incubated for 1 hour with ab10812 (1/1000). ab10812 clearly stained the nucleus (red). The cells were counterstained with DAPI (blue). 100x magnification.

  • ab10812 staining mouse embryonic stem cells by flow cytometry (gated on all living cells). The cells were incuabted with the antibody at 0.25ug/1.5 x 105 cells in a permeabilization buffer. A PE conjugated goat anti-rabbit antibody was used as the secondary.

    See Abreview

  • ab10812 staining Histone H3 (acetyl K9) in the root tips of Arabidopsis thaliana by Immunocytochemistry/ Immunofluorescence. Cells were fixed in 2% paraformaldehyde for 30 minutes at room temperature. Blocking and permeabilization was carried with 4% BSA solution containing 0,5% Triton X-100 in PBS at room temperature for 45 minutes. Slides were washed in PBS and incubated with primary antibody at a 1/200 dilution in 1% PBS for 1 hour at 37°C. Slides were washed in PBS and incubated with the secondary antibody ab6639 Goat polyclonal to Rabbit IgG - H&L (Cy3 ®) at a 1/500 dilution in 1% BSA in PBS for 1 hour at 37°C. Slides were counterstained with DAPI (2µg/mL) for 10 minutes at room temperature.
    Left image: DAPI stained interphase nucleus with prominent chromocenters
    Middle image: Distribution of Histone H3 (acetyl K9) in the nucleus (arrows indicate absence of signal from chromocenters)
    Right image: Merged image

    See Abreview

  • All lanes : Anti-Histone H3 (acetyl K9) antibody - ChIP Grade (ab10812) at 1/1000 dilution

    Lane 1 : Tissue lysate prepared from rat dorsal spinal cord at 5 µg
    Lane 2 : Tissue lysate prepared from rat dorsal spinal cord at 5 µg
    Lane 3 : Tissue lysate prepared from rat dorsal spinal cord at 8 µg
    Lane 4 : Tissue lysate prepared from rat dorsal spinal cord at 8 µg
    Lane 5 : Tissue lysate prepared from rat dorsal spinal cord at 8 µg with Human Histone H3 (acetyl K9) peptide (ab16635)

    Secondary
    HRP conjugated rabbit polyclonal
    developed using the ECL technique

    Predicted band size : 15 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)


    Exposure time : 2 minutes

    Image courtesy of an anonymous Abreview.

    Each sample is from a different extract.

    See Abreview

References for Anti-Histone H3 (acetyl K9) antibody - ChIP Grade (ab10812)

This product has been referenced in:
  • Conti Ad  et al. Dose- and time-dependent epigenetic changes in the livers of fisher 344 rats exposed to furan. Toxicol Sci 139:371-80 (2014). Read more (PubMed: 24614236) »
  • Hait NC  et al. Active, phosphorylated fingolimod inhibits histone deacetylases and facilitates fear extinction memory. Nat Neurosci 17:971-80 (2014). Mouse . Read more (PubMed: 24859201) »

See all 39 Publications for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing (12%)
Sample Zebrafish Tissue lysate - whole (Larvae (6dpf))
Specification Larvae (6dpf)
Treatment 10 uM Entinostat
Blocking step BSA as blocking agent for 16 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Username

Abcam user community

Verified customer

Submitted Sep 25 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Frozen sections)
Blocking step Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 24°C
Sample Mouse Tissue sections (thyroid tissue)
Specification thyroid tissue
Permeabilization Yes - 0.1% Tween 20
Fixative Paraformaldehyde
Username

Mr. Sanjay Gawade

Verified customer

Submitted Jun 26 2013

It may be worth trying ab10812 since each unit is 100ug, though we have not comopared it to ab4441 in any application. We do not have any more of ab4441 lot GR105501. We have several new lots but we do not test each lot in ChIP. The most recent data we...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Marmoset (common) Tissue sections (adult testis)
Specification adult testis
Fixative Formaldehyde
Antigen retrieval step Heat mediated - Buffer/Enzyme Used: Dako Antigen retrieval solution
Permeabilization No
Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Zachary Yu-Ching Lin

Verified customer

Submitted Oct 26 2012

Thank you very much for your call today and for letting us know about the trouble with ab4441 in Western blot.
As we discussed, I'm sending a free of charge vial of ab10812 on the order *** as well as a free of charge vial of the HeLa nuclear lysa...

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Thank you for contacting us.

Antibody ab113674 is the newest antibody of the three and is therefore less thoroughly characterized. Antibodies ab4441 and ab10812 have both been published many times and have received positive customer reviews ...

Read More

Thank you and your customer very much for your interest in ab10812.
To our knowledge, ab10812 has not been tested in IHC-Fr yet ,but is very likely to work in IHC-Fr since it is tested fro IHC-P. Therefore, I can offer a discount off a future purc...

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Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Rat Tissue lysate - other (dorsal spinal cord)
Loading amount 10 µg
Specification dorsal spinal cord
Gel Running Conditions Reduced Denaturing
Blocking step Milk as blocking agent for 45 minute(s) · Concentration: 5% · Temperature: 28°C
Username

Abcam user community

Verified customer

Submitted Feb 18 2012

Thank you for your enquiry.

Unfortunately we are not able to "save" specific vials of antibody for customer that may wish to purchase them in the future. If you'd like I can recommend specific lots that have the most stock, which ma...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Mouse Tissue lysate - whole (spinal cord)
Specification spinal cord
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: formaldehyde
Detection step Real-time PCR
Positive control total H3 content (ab1791 antibody), 10% input (not shown here)
Negative control gene desert primer pair, igg (not shown, but very low values)
Username

Abcam user community

Verified customer

Submitted Jan 06 2012

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"