RabMAb

Anti-Histone H3 (acetyl K56) antibody [EPR996Y] (ab76307)

Overview

  • Product nameAnti-Histone H3 (acetyl K56) antibody [EPR996Y]See all Histone H3 primary antibodies ...
  • Description
    Rabbit monoclonal [EPR996Y] to Histone H3 (acetyl K56)
  • Tested applicationsWB, IHC-P, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Saccharomyces cerevisiae, African Green Monkey
  • Immunogen

    A synthetic acetylated peptide corresponding to residues surrounding Lys 56 of Histone H3

  • Positive control
    • C6 cell lysate, untreated or treated with TSA; HeLa cells.
  • General notes

    Produced under U.S. Patent No. 5,675,063.

    This product is available conjugated to Alexa Fluor® 488 see ab170098.

    A 40 μl trial size is available to purchase for this antibody.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated ‘PUR’ on our product labels.

    If you have any questions regarding this update, please contact our Scientific Support team.

Properties

  • FormLiquid
  • Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage bufferPBS 49%,Sodium azide 0.01%,Glycerol 50%,BSA 0.05%
  • PurityTissue culture supernatant
  • Clonality Monoclonal
  • Clone numberEPR996Y
  • IsotypeIgG
  • Research Areas

Applications

Our Abpromise guarantee covers the use of ab76307 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
WB
IHC-P
ICC/IF
  • Application notesICC/IF: 1/100 - 1/250.
    IHC-P: 1/100 - 1/250. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. Use of HRP-conjugated or polymerized HRP secondary antibodies, stronger signals have been found using the polymerized HRP secondary.
    WB: 1/1000 - 1/2000. Detects a band of approximately 15 kDa (predicted molecular weight: 15 kDa).

    Is unsuitable for Flow Cyt or IP.


    Not yet tested in other applications.
    Optimal dilutions/concentrations should be determined by the end user.
  • 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 antibody
      • H3/A antibody
      • H31_HUMAN antibody
      • H3FA antibody
      • HIST1H3J antibody
      • Histone cluster 1, H3a 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 K56) antibody [EPR996Y] images

    • ab76307 staining Histone H3 (HEK56 Ac) in African Green Monkey kidney epithelial (CV-1 line) cell by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with Triton and blocked with 5% BSA for 1 hour at 21°C. Samples were incubated with primary antibody (1/500) for 16 hour at 4°C.

      CV-1 cells were infected with Simian virus 40 (SV40). 32 hours later the cells were incubated with EdU for 30 minutes before pre-extraction to detect active DNA replication foci and fixed, stained, and visualized by confocal microscopy.
      Blue - Hoechst stain, red - anti-T-Ag, magenta - EdU, green - anti-H3K56 Ac. Bar, 5 µm.

      See Abreview

    • ab76307 at 1/100 dilution staining Histone H3 in HeLa cells by Immunofluorescence.
    • All lanes : Anti-Histone H3 (acetyl K56) antibody [EPR996Y] (ab76307) at 1/2000 dilution

      Lane 1 : C6 cell lysate, untreated
      Lane 2 : C6 cell lysate, treated with TSA

      Lysates/proteins at 10 µg per lane.

      Secondary
      goat anti-rabbit HRP at 1/1000 dilution

      Predicted band size : 15 kDa
      Observed band size : 15 kDa
    • IHC image of ab76307 staining in Breast Cancer formalin fixed paraffin embedded tissue section, performed on a Leica 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 ab76307, undiluted, for 15 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.

      For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
    • ab76307 showing positive staining in Cervical carcinoma tissue.

    • ab76307 showing positive staining in Gastric adenocarcinoma tissue.

    • ab76307 showing positive staining in Lung adenocarcinoma tissue.

    • ab76307 showing positive staining in Lymphoma tissue.

    References for Anti-Histone H3 (acetyl K56) antibody [EPR996Y] (ab76307)

    This product has been referenced in:
    • Hagelkruys A  et al. A single allele of Hdac2 but not Hdac1 is sufficient for normal mouse brain development in the absence of its paralog. Development 141:604-16 (2014). WB, IHC-P ; Mouse . Read more (PubMed: 24449838) »
    • Sakakibara N  et al. Brd4 Is Displaced from HPV Replication Factories as They Expand and Amplify Viral DNA. PLoS Pathog 9:e1003777 (2013). ICC/IF ; Human . Read more (PubMed: 24278023) »

    See all 16 Publications for this product

    Product Wall

    Application Western blot
    Loading amount 20 µg
    Gel Running Conditions Reduced Denaturing
    Sample Xenopus laevis Cell lysate - other (Egg extracts and chromatin)
    Specification Egg extracts and chromatin
    Blocking step Milk as blocking agent for 30 minute(s) · Concentration: 5% · Temperature: 22°C
    Username

    Abcam user community

    Verified customer

    Submitted May 01 2014

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application Immunocytochemistry/ Immunofluorescence
    Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
    Sample African Green Monkey Cell (epithel kidney CV-1)
    Specification epithel kidney CV-1
    Permeabilization Yes - triton
    Fixative Paraformaldehyde
    Username

    Herr Dr. Gabor Rohaly

    Verified customer

    Submitted Jul 01 2013

    Thank you for contacting Abcam regarding ab76307.


    Below please find the IF protocol we suggest:


    Solutions and reagents

    1.1. Washing buffer:
    PBST washing buffer: 1X PBS / 0.1% Tween-20


    1.2. 2% Para...

    Read More

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