Anti-Histone H4 (acetyl K8) antibody - ChIP Grade (ab15823)

Overview

  • Product nameAnti-Histone H4 (acetyl K8) antibody - ChIP Grade
    See all Histone H4 primary antibodies
  • Description
    Rabbit polyclonal to Histone H4 (acetyl K8) - ChIP Grade
  • Tested applicationsSuitable for: ICC/IF, IHC-P, Electron Microscopy, ChIP, WBmore details
  • Species reactivity
    Reacts with: Mouse, Chicken, Cow, Human, Saccharomyces cerevisiae, Schizosaccharomyces pombe
  • Immunogen

    Synthetic peptide corresponding to Human Histone H4 aa 1-100 (internal sequence) (acetyl K8) conjugated to Keyhole Limpet Haemocyanin (KLH).
    Database link: P62805
    (Peptide available as ab15824)

  • Positive control
    • This antibody gave a positive signal on Hela Histone lysate-Butyrate treated and Calf Thymus Histone lysate.

Properties

Applications

Our Abpromise guarantee covers the use of ab15823 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
ICC/IF Use a concentration of 1 µg/ml.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Electron Microscopy Use at an assay dependent concentration. PubMed: 20543957
ChIP Use 2µl for 106 cells.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 12 kDa (predicted molecular weight: 11.2 kDa).

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 H4 family.
  • Post-translational
    modifications
    Acetylation at Lys-6 (H4K5ac), Lys-9 (H4K8ac), Lys-13 (H4K12ac) and Lys-17 (H4K16ac) occurs in coding regions of the genome but not in heterochromatin.
    Citrullination at Arg-4 (H4R3ci) by PADI4 impairs methylation.
    Monomethylation and asymmetric dimethylation at Arg-4 (H4R3me1 and H4R3me2a, respectively) by PRMT1 favors acetylation at Lys-9 (H4K8ac) and Lys-13 (H4K12ac). Demethylation is performed by JMJD6. Symmetric dimethylation on Arg-4 (H4R3me2s) by the PRDM1/PRMT5 complex may play a crucial role in the germ-cell lineage.
    Monomethylated, dimethylated or trimethylated at Lys-21 (H4K20me1, H4K20me2, H4K20me3). Monomethylation is performed by SET8. Trimethylation is performed by SUV420H1 and SUV420H2 and induces gene silencing.
    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. Monoubiquitinated at Lys-92 of histone H4 (H4K91ub1) in response to DNA damage. The exact role of H4K91ub1 in DNA damage response is still unclear but it may function as a licensing signal for additional histone H4 post-translational modifications such as H4 Lys-21 methylation (H4K20me).
    Sumoylated, which is associated with transcriptional repression.
  • Cellular localizationNucleus. Chromosome.
  • Information by UniProt
  • Database links
    see all
  • Alternative names
    • Histone gene cluster 1, H4A antibody
    • Histone gene cluster 2, H4 antibody
    • dJ160A22.1 antibody
    • dJ160A22.2 antibody
    • dJ221C16.1 antibody
    • dJ221C16.9 antibody
    • FO108 antibody
    • H4 antibody
    • H4 histone family, member A antibody
    • H4 histone family, member B antibody
    • H4 histone family, member C antibody
    • H4 histone family, member D antibody
    • H4 histone family, member E antibody
    • H4 histone family, member G antibody
    • H4 histone family, member H antibody
    • H4 histone family, member I antibody
    • H4 histone family, member J antibody
    • H4 histone family, member K antibody
    • H4 histone family, member M antibody
    • H4 histone family, member N antibody
    • H4 histone, family 2 antibody
    • H4/A antibody
    • H4/B antibody
    • H4/C antibody
    • H4/D antibody
    • H4/E antibody
    • H4/G antibody
    • H4/H antibody
    • H4/I antibody
    • H4/J antibody
    • H4/K antibody
    • H4/M antibody
    • H4/N antibody
    • H4/O antibody
    • H4/p antibody
    • H4_HUMAN antibody
    • H4F2 antibody
    • H4F2iii antibody
    • H4F2iv antibody
    • H4FA antibody
    • H4FB antibody
    • H4FC antibody
    • H4FD antibody
    • H4FE antibody
    • H4FG antibody
    • H4FH antibody
    • H4FI antibody
    • H4FJ antibody
    • H4FK antibody
    • HIST1 cluster, H4A antibody
    • HIST1 cluster, H4B antibody
    • HIST1 cluster, H4D antibody
    • HIST2H4 antibody
    • Hist4 cluster, H4 antibody
    • Hist4h4 antibody
    • histone 1, H4a antibody
    • histone 1, H4c antibody
    • histone 1, H4d antibody
    • histone 1, H4f antibody
    • histone 1, H4h antibody
    • histone 1, H4i antibody
    • histone 1, H4j antibody
    • histone 1, H4k antibody
    • histone 1, H4l antibody
    • histone 2, H4a antibody
    • histone 2, H4b antibody
    • Histone 4 family, member M antibody
    • histone 4, H4 antibody
    • histone cluster 1, H4 antibody
    • histone cluster 1, H4a antibody
    • histone cluster 1, H4b antibody
    • histone cluster 1, H4c antibody
    • histone cluster 1, H4d antibody
    • histone cluster 1, H4e antibody
    • histone cluster 1, H4f antibody
    • histone cluster 1, H4h antibody
    • histone cluster 1, H4i antibody
    • histone cluster 1, H4j antibody
    • histone cluster 1, H4k antibody
    • histone cluster 1, H4l antibody
    • histone cluster 2, H4a antibody
    • histone cluster 2, H4b antibody
    • histone cluster 4, H4 antibody
    • Histone family, member A antibody
    • Histone family, member B antibody
    • Histone family, member D antibody
    • Histone family, member H antibody
    • Histone family, member I antibody
    • Histone family, member L antibody
    • Histone gene cluster 1, H4 antibody
    • Histone gene cluster 1, H4D antibody
    • Histone gene cluster 1, H4E antibody
    • Histone gene cluster 1, H4K antibody
    • Histone gene cluster 4, H4 antibody
    • Histone gene cluster 4, H4 histone antibody
    • Histone H4 antibody
    • histone IV, family 2 antibody
    see all

Anti-Histone H4 (acetyl K8) antibody - ChIP Grade images

  • IHC image of Histone H4 (acetyl K8) 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 ab15823, 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.
  • Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 min. The  ChIP was performed with 25 µg of chromatin, 2 µg of ab15823 (blue), and 20 µl 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.

  • All lanes : Anti-Histone H4 (acetyl K8) antibody - ChIP Grade (ab15823) at 1 µg/ml

    Lane 1 : Sodium butyrate treated hela histone
    Lane 2 : Untreated hela histone
    Lane 3 : Sodium butyrate treated hela histone with Human Histone H4 (acetyl K8) peptide (ab15824) at 1 µg/ml
    Lane 4 : Untreated hela histone with Human Histone H4 (acetyl K8) peptide (ab15824) at 1 µg/ml
    Lane 5 : Sodium butyrate treated hela histone with Human Histone H4 peptide (ab15825) at 1 µg/ml
    Lane 6 : Untreated hela histone with Human Histone H4 peptide (ab15825) at 1 µg/ml
    Lane 7 : Sodium butyrate treated hela histone with Histone H4 peptide - acetyl K5 (ab15596) at 1 µg/ml
    Lane 8 : Untreated hela histone with Histone H4 peptide - acetyl K5 (ab15596) at 1 µg/ml
    Lane 9 : Sodium butyrate treated hela histone with Human Histone H4 (acetyl K12) peptide (ab15662) at 1 µg/ml
    Lane 10 : Untreated hela histone with Human Histone H4 (acetyl K12) peptide (ab15662) at 1 µg/ml
    Lane 11 : Sodium butyrate treated hela histone with Histone H2A peptide - acetyl K9 at 1 µg/ml
    Lane 12 : Untreated hela histone with Histone H2A peptide - acetyl K9 at 1 µg/ml

    Lysates/proteins at 2.5 µg per lane.

    Secondary
    IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 11.2 kDa
    Observed band size : 12 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 22 kDa (possible cross reactivity).

    The blot shows that a H4 peptide containing an acetylated K8 residue was able to block recognition of histones by ab15823. Peptides containing other acetylated K residues of H4 and H2A did not block binding of histones by ab15823. This suggests ab15823 specifically recognises Histone H4 acetyl K8.

  • ICC/IF image of ab15823 stained human HeLa cells. The cells were methanol fixed (5 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab15823, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. 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). DAPI was used to stain the cell nuclei (blue).

  • A: Overall view of a murine rod photoreceptor nucleus immunogold labelled with acetylated H4K8-specific antibodies.
    B: Schematic representation of the H4K8ac-specific labelling. The bar represents 1µm.
  • All lanes : Anti-Histone H4 (acetyl K8) antibody - ChIP Grade (ab15823) at 1 µg/ml

    Lane 1 : Calf Thymus Histone Preparation Nuclear Lysate (ab121)
    Lane 2 : Calf Thymus Histone Preparation Nuclear Lysate (ab121) with Human Histone H4 peptide (ab15825) at 0.5 µg/ml
    Lane 3 : Calf Thymus Histone Preparation Nuclear Lysate (ab121) with Human Histone H4 (acetyl K8) peptide (ab15824) at 0.5 µg/ml
    Lane 4 : Calf Thymus Histone Preparation Nuclear Lysate (ab121) with Histone H4 peptide - acetyl K5 at 0.5 µg/ml
    Lane 5 : Calf Thymus Histone Preparation Nuclear Lysate (ab121) with Human Histone H4 (acetyl K12) peptide (ab15662) at 0.5 µg/ml
    Lane 6 : Calf Thymus Histone Preparation Nuclear Lysate (ab121) with Human Histone H2A (acetyl K9) peptide (ab54016) at 0.5 µg/ml

    Lysates/proteins at 0.5 µg/ml per lane.

    Secondary
    Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size : 11.2 kDa
    Observed band size : 13 kDa (why is the actual band size different from the predicted?)


    Exposure time : 3 minutes

References for Anti-Histone H4 (acetyl K8) antibody - ChIP Grade (ab15823)

This product has been referenced in:
  • Banerjee A  et al. The DNA intercalators ethidium bromide and propidium iodide also bind to core histones. FEBS Open Bio 4:251-9 (2014). WB ; Human . Read more (PubMed: 24649406) »
  • Azad GK  et al. Assessment of the biological pathways targeted by isocyanate using N-succinimidyl N-methylcarbamate in budding yeast Saccharomyces cerevisiae. PLoS One 9:e92993 (2014). WB ; Chicken . Read more (PubMed: 24664350) »

See all 19 Publications for this product

Product Wall

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing (12)
Sample Human Cell lysate - whole cell (Human HeLa cells)
Specification Human HeLa cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 23°C
Username

Abcam user community

Verified customer

Submitted Nov 13 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (retina)
Specification retina
Fixative 2.5% PAF + 0.1% Glu
Permeabilization No
Blocking step BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Caroline Kizilyaprak

Verified customer

Submitted Aug 26 2010

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Mouse Cell lysate - whole cell (skin epidermis)
Specification skin epidermis
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% formaldehyde
Detection step Semiquantitative PCR
Negative control rabbit IgG immunoprecipitated sample did not show significant PCR amplification.
Username

Abcam user community

Verified customer

Submitted Jan 25 2008

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