Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)
Overview
- Product nameAnti-Histone H3 (acetyl K27) antibody - ChIP GradeSee all Histone H3 primary antibodies ...
- DescriptionRabbit polyclonal to Histone H3 (acetyl K27) - ChIP Grade
- Specificityab4729 recognises acetyl K27 histone H3 in western blot and immunofluorescence. However, it may also recognise to a small degree acetyl K9 histone H3 (immunofluorescence) and unmodified histone H3 (western blot). See figures below.
- Tested applicationsIHC-Fr, ICC/IF, WB, IHC-P, CHIPseq, ChIP/Chip, ChIP more details
- Species reactivityReacts with: Mouse, Rat, Cow, Human, Arabidopsis thaliana, Fruit fly (Drosophila melanogaster), Monkey, Plasmodium falciparum, Cyanidioschyzon merolae
Predicted to work with: Xenopus laevis, Caenorhabditis elegans - Immunogen
Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, acetylated at K27.
(Peptide available as ab24404.)
- Positive controlCalf thymus histone lysate; HeLa whole cell lysate; HeLa nuclear lysate; HeLa histone prep Butyrate treated.
Properties
- FormLiquid
- Storage instructionsStore at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
- Storage bufferPreservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS. pH 7.4 -
Concentration information loading... - PurityImmunogen affinity purified
- Clonality Polyclonal
- IsotypeIgG
- Research Areas
Applications
Our Abpromise guarantee covers the use of ab4729 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
| Application | Notes |
|---|---|
| IHC-Fr | IHC-Fr: Use at an assay dependent concentration. |
| ICC/IF | ICC/IF: 1/500. Can be used with paraformaldehyde- or methanol- fixed cells. |
| WB | WB: 1/1000. Detects a band of approximately 17 kDa.Can be blocked with Histone H3 peptide - acetyl K27 (ab24404). |
| IHC-P | IHC-P: Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. |
| CHIPseq | CHIPseq: Use at an assay dependent dilution. |
| ChIP/Chip | ChIP/Chip: Use at an assay dependent dilution. |
| ChIP | ChIP: 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
modificationsAcetylation 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.
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Database links
- Entrez Gene: 8350 Human
- Entrez Gene: 8351 Human
- Entrez Gene: 8352 Human
- Entrez Gene: 8353 Human
- Entrez Gene: 8354 Human
- Entrez Gene: 8355 Human
- Entrez Gene: 8356 Human
- Entrez Gene: 8357 Human
- Entrez Gene: 8358 Human
- Entrez Gene: 8968 Human
- Entrez Gene: 319152 Mouse
- Entrez Gene: 319153 Mouse
- Entrez Gene: 360198 Mouse
- Entrez Gene: 97908 Mouse
- Entrez Gene: 100364501 Rat
- Entrez Gene: 100365669 Rat
- Entrez Gene: 291159 Rat
- Entrez Gene: 679994 Rat
- Entrez Gene: 680511 Rat
- Entrez Gene: 682330 Rat
- Omim: 601128 Human
- Omim: 142780 Human
- Omim: 601058 Human
- SwissProt: P84243 Human
- SwissProt: Q71DI3 Human
- SwissProt: P68431 Human
- SwissProt: P68433 Mouse
- SwissProt: Q6LED0 Rat
- Unigene: 132854 Human
- Unigene: 247813 Human
- Unigene: 247814 Human
- Unigene: 248176 Human
- Unigene: 443021 Human
- Unigene: 484990 Human
- Unigene: 532144 Human
- Unigene: 533292 Human
- Unigene: 546315 Human
- Unigene: 586261 Human
- Unigene: 591778 Human
- Unigene: 221301 Mouse
- Unigene: 261657 Mouse
- Unigene: 377874 Mouse
- Unigene: 390558 Mouse
- Unigene: 397328 Mouse
- Unigene: 138090 Rat
see all
Target information above from: UniProt accession
P68431
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010)
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Alternative names
- acetyl H3 antibodyH 3 antibodyH3 3 Like Sequence MH921 antibody
- H3 3A antibodyH3 antibodyH3 Histone antibodyH3 Histone Family Member E Pseudogene antibodyH3.4 antibodyH3/A antibodyH3/g antibodyH31_HUMAN antibodyH3F3 antibodyH3FA antibodyH3FT antibodyH3t antibodyHIST1H3J antibodyHIST3H3 antibodyHistone cluster 1, H3a antibodyHistone H3 3 Pseudogene antibodyHistone H3.1 antibodyHistone H3.3 antibodyHistone H3/a antibodyHistone H3/b antibodyHistone H3/c antibodyHistone H3/d antibodyHistone H3/f antibodyHistone H3/h antibodyHistone H3/i antibodyHistone H3/j antibodyHistone H3/k antibodyHistone H3/l antibody
see all
Anti-Histone H3 (acetyl K27) antibody - ChIP Grade images
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ChIP - Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)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 ab4729 (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.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)IHC image of Histone H3 (acetyl K27) 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 ab4729, 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. -
Western blot - Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)All lanes : Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729) at 1 µg/ml
Lane 1 : HeLa Histone Preparation Nuclear Lysate - Butyrated
Lane 2 : HeLa Histone Preparation Nuclear Lysate - Butyrated with Histone H3 peptide - unmodified R17 at 0.5 µg/ml
Lane 3 : HeLa Histone Preparation Nuclear Lysate - Butyrated withHistone H3 peptide - acetyl K9 (ab16635) at 0.5 µg/ml
Lane 4 : HeLa Histone Preparation Nuclear Lysate - Butyrated with Histone H3 peptide - acetyl K14 at 0.5 µg/ml
Lane 5 : HeLa Histone Preparation Nuclear Lysate - Butyrated withHistone H3 peptide - acetyl K18 (ab24003) at 0.5 µg/ml
Lane 6 : HeLa Histone Preparation Nuclear Lysate - Butyrated withHistone H3 peptide - acetyl K23 (ab48359) at 0.5 µg/ml
Lane 7 : HeLa Histone Preparation Nuclear Lysate - Butyrated withHistone H3 peptide - acetyl K27 (ab24404) at 0.5 µg/ml
Lane 8 : HeLa Histone Preparation Nuclear Lysate - Butyrated with Histone H3 peptide - acetyl K36 at 0.5 µg/ml
Lane 9 : HeLa Histone Preparation Nuclear Lysate - Butyrated withHistone H4 peptide - acetyl K12 (ab15662) at 0.5 µg/ml
Lysates/proteins at 2.5 µg per lane.
Secondary
Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Performed under reducing conditions.
Predicted band size : 15.4 kDa
Observed band size : 17 kDa (why is the actual band size different from the predicted?)
Additional bands at : 12 kDa. We are unsure as to the identity of these extra bands.
Exposure time : 150 seconds -
Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)Image courtesy of an Abreview submitted by Dr. Kirk McManus, Univ. of Manitoba/Cancer Care MICB, Canadaab4729 (1/500) staining Histone H3 (acetyl K27) in HeLa cells (green). Cells were fixed in paraformaldehyde, permeabilized with 0.5% Triton X-100/PBS and counterstained with DAPI in order to highlight the nucleus (red). For further experimental details please see abreview. -
Immunofluorescence - Histone H3 (acetyl K27) antibody (ab4729)Petra Hajkova - Gurdon Institute, Cambridge UniversityPrimary antibody: ab4729 (H3 acetyl K27)
Dilution: 1/100ab4729 strongly stained histones of mouse ES cells. However, fluroescence was greatly diminished following pre-blocking using a H3 acetyl K9 peptide. This suggests the antibody cross-reacts with the K9 and K27 residues.
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Western blot - Histone H3 (acetyl K27) antibody (ab4729)Lanes 1 & 3 : Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729) at 0.2 µg/ml
Lanes 2 & 4 : Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729) at 0.1 µg/ml
Lane 1 : Calf thymus histone lysate
Lane 2 : Calf thymus histone lysate
Lane 3 : Calf thymus histone lysate withHistone H3 peptide - acetyl K27 (ab24404) at 2 µg
Lane 4 : Calf thymus histone lysate withHistone H3 peptide - acetyl K27 (ab24404) at 2 µg
Lysates/proteins at 1 µg per lane.
Secondary
Goat anti-rabbit (HRP) at 1/2000 dilution
Predicted band size : 15.4 kDa
Observed band size : 17 kDa (why is the actual band size different from the predicted?) -
Western blot - Histone H3 (acetyl K27) antibody (ab4729)All lanes : Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729) at 0.2 µg/ml
Lane 1 : HeLa whole cell lysate
Lane 2 : HeLa nuclear lysate
Lane 3 : HeLa whole cell lysate withHistone H3 peptide (ab17163) at 1 µg/ml
Lane 4 : HeLa nuclear lysate withHistone H3 peptide (ab17163) at 1 µg/ml
Lane 5 : HeLa whole cell lysate withHistone H3 peptide - acetyl K27 (ab24404) at 1 µg/ml
Lane 6 : HeLa nuclear lysate withHistone H3 peptide - acetyl K27 (ab24404) at 1 µg/ml
Lane 7 : HeLa whole cell lysate withHistone H3 peptide - acetyl K9 (ab16635) at 1 µg/ml
Lane 8 : HeLa nuclear lysate withHistone H3 peptide - acetyl K9 (ab16635) at 1 µg/ml
Lane 9 : HeLa whole cell lysate withHistone H3 peptide - acetyl K18 (ab24003) at 1 µg/ml
Lane 10 : HeLa nuclear lysate withHistone H3 peptide - acetyl K18 (ab24003) at 1 µg/ml
Lane 11 : HeLa whole cell lysate withHistone H3 peptide - asymmetric di methyl R26 (ab2854) at 1 µg/ml
Lane 12 : HeLa nuclear lysate withHistone H3 peptide - asymmetric di methyl R26 (ab2854) at 1 µg/ml
Lane 13 : HeLa whole cell lysate withHistone H4 peptide - acetyl K12 (ab15662) at 1 µg/ml
Lane 14 : HeLa nuclear lysate withHistone H4 peptide - acetyl K12 (ab15662) at 1 µg/ml
Lysates/proteins at 20 µg per lane.
Secondary
Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP) (ab6721) at 1/5000 dilution
Predicted band size : 15.4 kDa
Observed band size : 17 kDa (why is the actual band size different from the predicted?) -
Immunocytochemistry/ Immunofluorescence - Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)ICC/IF image of ab4729 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 (ab4729, 1µg/ml) overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti-rabbit IgG - H&L, pre-adsorbed (ab96899) used at a 1/250 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. -
Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)This image is courtesy of an Abreview submitted by Toshiyuki Soneab4729 staining Histone H3 (acetyl K27) in Cyanidioschyzon merolae cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde and blocked with 5% BSA for 30 minutes at 37°C. Samples were incubated with primary antibody (1/100 in PBS + 0.1% BSA) for 1 hour at 37°C. An Alexa Fluor®488-conjugated Goat anti-rabbit polyclonal (100)was used as the secondary antibody.
References for Anti-Histone H3 (acetyl K27) antibody - ChIP Grade (ab4729)
This product has been referenced in:
- Lau PN & Cheung P Elucidating combinatorial histone modifications and crosstalks by coupling histone-modifying enzyme with biotin ligase activity. Nucleic Acids Res 41:e49 (2013). WB ; Human . Read more (PubMed: 23258705) »
- Visser M et al. HERC2 rs12913832 modulates human pigmentation by attenuating chromatin-loop formation between a long-range enhancer and the OCA2 promoter. Genome Res 22:446-55 (2012). ChIP . Read more (PubMed: 22234890) »
