Anti-Histone H3.3 antibody (ab94817)

Overview

  • Product nameAnti-Histone H3.3 antibody
    See all Histone H3.3 primary antibodies
  • Description
    Rabbit polyclonal to Histone H3.3
  • SpecificityThis antibody was raised against a peptide derived from within residues 50 to the C-terminus of Human Histone H3.3. However, due to the extensive amount of homology between H3 variants, we expect this antibody to show some cross reactivity with H3.1 and H3.2.
  • Tested applicationsSuitable for: WBmore details
  • Species reactivity
    Reacts with: Mouse, Human
    Predicted to work with: Rat, Rabbit, Chicken, Cow, Pig, Zebrafish, Orangutan
  • Immunogen

    Synthetic peptide corresponding to Human Histone H3.3 aa 50 to the C-terminus conjugated to Keyhole Limpet Haemocyanin (KLH).
    Database link: P84243
    (Peptide available as ab129219)

  • Positive control
    • This antibody gave a positive signal in the following whole cell lysates: HeLa; SAOS-2; Jurkat; NIH3T3.

Properties

  • FormLiquid
  • Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage bufferpH: 7.40
    Preservative: 0.02% Sodium azide
    Constituent: PBS
    Note: Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
  • Concentration information loading...
  • PurityImmunogen affinity purified
  • ClonalityPolyclonal
  • IsotypeIgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab94817 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 Use a concentration of 1 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa).

Target

  • FunctionVariant histone H3 which replaces conventional H3 in a wide range of nucleosomes in active genes. Constitutes the predominant form of histone H3 in non-dividing cells and is incorporated into chromatin independently of DNA synthesis. Deposited at sites of nucleosomal displacement throughout transcribed genes, suggesting that it represents an epigenetic imprint of transcriptionally active chromatin. 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 throughout the cell cycle independently of DNA synthesis.
  • 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.
    Specifically enriched in modifications associated with active chromatin such as methylation at Lys-5 (H3K4me), Lys-37 and Lys-80. 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), which are linked to gene repression, are underrepresented. 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. Phosphorylation on Ser-32 (H3S31ph) is specific to regions bordering centromeres in metaphase chromosomes.
    Ubiquitinated. Monoubiquitinated by RAG1 in lymphoid cells, monoubiquitination is required for V(D)J recombination.
  • Cellular localizationNucleus. Chromosome.
  • Information by UniProt
  • Database links
  • Alternative names
    • H3 histone family 3A antibody
    • H3 histone family 3B antibody
    • H3 histone, family 3B (H3.3B) antibody
    • H3.3 antibody
    • H3.3A antibody
    • H3.3B antibody
    • H33_HUMAN antibody
    • H3F3 antibody
    • H3F3A antibody
    • H3f3b antibody
    • Histone H3.3 antibody
    • Histone H3.3Q antibody
    • Histone H3.A antibody
    • Histone H3.B antibody
    • MGC87782 antibody
    • MGC87783 antibody
    see all

Anti-Histone H3.3 antibody images

  • All lanes : Anti-Histone H3.3 antibody (ab94817) at 1 µg/ml

    Lane 1 : Calf Thymus Histone Preparation Nuclear Lysate at 0.5 µg
    Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 20 µg
    Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 20 µg
    Lane 4 : Histone H3.1 Recombinant Protein at 0.1 µg
    Lane 5 : Histone H3.3 Recombinant Protein at 0.1 µg

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/10000 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 : 20 minutes
  • All lanes : Anti-Histone H3.3 antibody (ab94817) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : HeLa Histone Preparation Nuclear Lysate
    Lane 3 : Saos 2 (Human epithelial-like osteosarcoma cell line) Whole Cell Lysate
    Lane 4 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 5 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Histone H3.3 peptide (ab129219) at 1 µg/ml
    Lane 6 : HeLa Histone Preparation Nuclear Lysate with Histone H3.3 peptide (ab129219) at 1 µg/ml
    Lane 7 : Saos 2 (Human epithelial-like osteosarcoma cell line) Whole Cell Lysate with Histone H3.3 peptide (ab129219) at 1 µg/ml
    Lane 8 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate with Histone H3.3 peptide (ab129219) at 1 µg/ml
    Lane 9 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Histone H3.1 Peptide at 1 µg/ml
    Lane 10 : HeLa Histone Preparation Nuclear Lysate with Histone H3.1 Peptide at 1 µg/ml
    Lane 11 : Saos 2 (Human epithelial-like osteosarcoma cell line) Whole Cell Lysate with Histone H3.1 Peptide at 1 µg/ml
    Lane 12 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate with Histone H3.1 Peptide at 1 µg/ml

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
    Developed using the ECL technique

    Performed under reducing conditions.

    Predicted band size : 15 kDa


    Exposure time : 12 minutes
  • All lanes : Anti-Histone H3.3 antibody (ab94817) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : Saos 2 (Human epithelial-like osteosarcoma cell line) Whole Cell Lysate
    Lane 3 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate
    Lane 4 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) 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?)
    Additional bands at : 100 kDa,50 kDa,70 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 8 minutes

References for Anti-Histone H3.3 antibody (ab94817)

ab94817 has not yet been referenced specifically in any publications.

Product Wall

Abcam has not validated the combination of species/application used in this Abreview.
Application Western blot
Sample Zebrafish Cell lysate - whole cell (embryo, sphere stage (plus 8cell - adult stage))
Gel Running Conditions Reduced Denaturing (NuPAGE Bis-Tris gel 4-12%)
Loading amount 20000 cells
Specification embryo, sphere stage (plus 8cell - adult stage)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Jun 25 2013

Application Western blot
Sample Mouse Cell lysate - other (Histones (acid extracted from NIH 3T3 cells))
Loading amount 50000 cells
Specification Histones (acid extracted from NIH 3T3 cells)
Gel Running Conditions Reduced Denaturing (AUT gel)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C
Username

Abcam user community

Verified customer

Submitted Oct 31 2012

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement for one vial of ab94817.

To check the status of the order please contact o...

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