Anti-Histone H3 抗体 - Nuclear Loading Control and ChIP Grade (ab1791)

製品の概要

  • 製品名Anti-Histone H3 antibody - Nuclear Loading Control and ChIP Grade
    Histone H3 一次抗体 製品一覧
  • 製品の詳細
    Rabbit polyclonal to Histone H3 - Nuclear Loading Control and ChIP Grade
  • アプリケーション適用あり: IHC-Fr, CHIPseq, Dot blot, Flow Cyt, IHC-P, Electron Microscopy, ICC/IF, ChIP, IP, WB, ChIP/Chip, IHC - Wholemount, ICCmore details
  • 種交差性
    交差種: Mouse, Rat, Chicken, Dog, Human, Saccharomyces cerevisiae, Xenopus laevis, Arabidopsis thaliana, Caenorhabditis elegans, Drosophila melanogaster, Ferret, Indian Muntjac, Schizosaccharomyces pombe, Zebrafish, Silk worm, Rainbow Trout, Trypanosoma cruzi, Neurospora crassa , Toxoplasma gondii, Rice, Schistosoma mansoni , Candida albicans, Cyanidioschyzon merolae
    交差が予測される動物種: a wide range of other species, all Mammals
  • 免疫原

    Synthetic peptide corresponding to Human Histone H3 aa 100 to the C-terminus conjugated to Keyhole Limpet Haemocyanin (KLH).
    Database link: P68431
    (Peptide available as ab12149)

  • ポジティブ・コントロール
    • WB: HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate. Available to buy from Abcam under ab150035. Drosophila embryo nuclear extract, NIH3t3, S.cerevisiae (Y190) and S.pombe Whole Cell Lysates. ICC/IF: Methanol fixed HeLa cells. ChIP: Chromatin from HeLa cells.
  • 特記事項

    ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab1791 in the following tested applications.

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

アプリケーション Abreviews 特記事項
IHC-Fr 1/200.
CHIPseq Use at an assay dependent concentration.
Dot blot 1/10000.
Flow Cyt Use at an assay dependent concentration.

ab171870 - Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.

IHC-P 1/100 - 1/400. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Electron Microscopy 1/50. Customer feedback
ICC/IF Use a concentration of 1 µg/ml.

Methanol fixed cells.

 

ChIP Use 2µg for 106 cells.
IP Use a concentration of 5 µg/ml.
WB 1/1000 - 1/5000. Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa).Can be blocked with Human Histone H3 peptide (ab12149).
ChIP/Chip Use at an assay dependent concentration.
IHC - Wholemount Use at an assay dependent concentration. PubMed: 22219645
ICC 1/100 - 1/500.

ターゲット情報

  • 機能Core 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.
  • 配列類似性Belongs to the histone H3 family.
  • 発生段階Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
  • 翻訳後修飾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.
  • 細胞内局在Nucleus. Chromosome.
  • Information by UniProt
  • 参照データベース
  • 別名
    • H3 histone family, member A antibody
    • H3/A antibody
    • H31_HUMAN antibody
    • H3FA antibody
    • Hist1h3a antibody
    • HIST1H3B antibody
    • HIST1H3C antibody
    • HIST1H3D antibody
    • HIST1H3E antibody
    • HIST1H3F antibody
    • HIST1H3G antibody
    • HIST1H3H antibody
    • HIST1H3I antibody
    • HIST1H3J antibody
    • histone 1, H3a antibody
    • Histone cluster 1, H3a antibody
    • Histone H3.1 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 antibody - Nuclear Loading Control and ChIP Grade 画像

  • ab1791 staining Histone H3 in HeLa by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with methanol and blocked with 0.2% fish scale gelatin for 1 hour at 25°C. Samples were incubated with the primary antibody (1/300 in PBS + 0.2% gelatin) for 20 minutes at 25°C. An Alexa Fluor® 488-conjugated donkey anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.

    Green - Histone H3.
    Blue - DAPI.
    Red - Tubulin.

    See Abreview

  • Chromatin from Xenopus laevis oocytes was prepared according to the Abcam X-ChIP protocol. Oocytes were fixed with formaldehyde for 10 min. The ChIP was performed with 25 mg of chromatin, 3 mg of ab7834 (anti-H3, light blue) and 3 µg of ab1791 (anti-H3, dark blue), and 20 ml of Protein A/G sepharose beads. A non-specific antibody was used as a control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach).
  • All lanes : Anti-Histone H3 antibody - Nuclear Loading Control and ChIP Grade (ab1791) at 1/1000 dilution

    Lane 1 : A431 (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate
    Lane 3 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate
    Lane 4 : A431 (Human epithelial carcinoma cell line) Whole Cell Lysate with Human Histone H3 peptide (ab12149) at 1 µg/ml
    Lane 5 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate with Human Histone H3 peptide (ab12149) at 1 µg/ml
    Lane 6 : HEK293 (Human embryonic kidney cell line) Whole Cell Lysate with Human Histone H3 peptide (ab12149) at 1 µg/ml

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab6721) 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 : 10 seconds

    This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab1791 overnight at 4°C. Antibody binding was visualised using ECL development solution ab133406.

  • ab1796 staining Histone H3 in mouse liver tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde, permeabilized with 0.05% Triton X-100 in PBS for 30 minutes and blocked with 5% BSA for 1 hour; antigen retrieval was by heat mediation in sodium citrate pH 6. Samples were incubated with the primary antibody (1/500 in blocking buffer) for 16 hours at 4°C. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG polyclonal (1/400) was used as the secondary antibody.

    See Abreview

  • Histone H3 - ChIP Grade was immunoprecipitated using 0.5mg HeLa whole cell extract, 5µg of Rabbit polyclonal to and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).

    The antibody was incubated under agitation with Protein G beads for 10min, HeLa whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.

    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab1791.

    Secondary Antibody: Mouse anti-rabbit HRP light chain (HRP) (ab99697).

    Band: 15kDa; Histone H3 - ChIP Grade



  • Predicted band size : 15 kDa

    This image is courtesy of John E. Mueller and J. Ruth German (Mary Bryk lab)

    Rabbit polyclonal to Histone H3 (ab1791) at 1/5000 on S. cerevisiae whole cell lysate (40 ug per lane).

    Protein resolved on 15% SDS-PAGE gel. After transfer to PVDF membrane, blots were blocked in 1X PBS, 0.1% Tween-20, and 5% milk. ab1791 was diluted in 5 ml blocking buffer at 1/5000.  Blots plus primary antibodies were either incubated overnight at 4C or at RT for 2 hr. Blots were washed 6X for 10 min each in PBS with 0.1% Tween-20 before addition of secondary antibodies. Secondary antibodies were diluted 1/2,000 in blocking buffer and incubated with blots for 2 hr at RT. Secondary blots were washed 4X for 10 min each in PBS with 0.1% Tween-20 and 2X for 10 min each in PBS.
  • ab1791 staining Histone H3 in Human infantile fibromatosis tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% FBS/BSA for 3 hours at room temperature; antigen retrieval was by heat mediation in Tris pH9. Samples were incubated with primary antibody (1/100 in TBS + 1% BSA + 1% FBS) for 16 hours. An undiluted HRP-conjugated goat anti-rabbit IgG polyclonal was used as the secondary antibody.

    See Abreview

  • All lanes : Anti-Histone H3 antibody - Nuclear Loading Control and ChIP Grade (ab1791) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : NIH 3T3 whole cell lysate (ab7179)
    Lane 3 : Drosophila embryo nuclear extract (from melanogaster embryos 0-12Hr)
    Lane 4 : S.cerevisiae (Y190) Whole Cell Lysate
    Lane 5 : S.pombe Whole Cell Lysate

    Lysates/proteins at 10 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 kDa
    Observed band size : 17 kDa (why is the actual band size different from the predicted?)
    ab1791 is tested in western blot on a range of species.  We recommend loading higher amounts of protein (20-30ug) to increase the signal in yeast lysates
  • ab1791 staining Histone H3 (red) in rat brain tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde, permeabilized with 0.1% TBS-TritionX and blocked with 10% serum for 1 hour at 25°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with the primary antibody (1/500 in 10% normal goat serum) for 24 hours at 24°C. An Alexa Fluor® 594-conjugated goat anti-rabbit IgG polyclonal (1/500) was used as the secondary antibody.

    Green - Nucleus staining.
    Red - Histone H3 staining.

  • All lanes : Anti-Histone H3 antibody - Nuclear Loading Control and ChIP Grade (ab1791) at 1/1000 dilution

    Lane 1 : Mouse skeletal muscle mitochondrial fraction
    Lane 2 : Mouse skeletal muscle mitochondrial fraction

    Lysates/proteins at 20 µg per lane.

    Secondary
    HRP-conjugated goat anti-rabbit IgG at 1/4000 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 : 7 minutes

    This image is courtesy of an anonymous Abreview

    Blocked with 3% milk for 1 hour at 25°C.

    Incubated with the primary antibody for 16 hours at 4°C in 3% milk in TBS-tween.

    See Abreview

  • Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab1791 (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 for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

  •  ab1791 staining mouse embryonic stem cells by flow cytometry (gated on all living cells). The cell colonies were trypsinized and incubated with the antibody 1ug/1.5 x 105cells in a permeabilization buffer. A PE conjugated goat anti-rabbit antibody was used as the secondary.

    See Abreview

  • Histone H3 immunogold detection in HeLa cells.
    Ulltra-thin sections of paraformaldehyde-fixed, Lowicryl K4M embedded cells were incubated sequentially with ab1791 antibody and an anti-rabbit antibody coupled to 10 nm gold particles. Notice absence of chromatin within Interchromatin Granules (IG), a reservoir of spliceosomal snRNPs, and high labeling of patches of condensed chromatin close to the nuclear envelope (arrows). Nu: nucleus, Cyt: cytoplasm.

    Image courtesy of Gerard Pierron, IGR-Villejuif, France.

    Sample : human
    Type : HeLa cells
    Fixative : either paraformaldehyde 4% or 1.6% glutaraldehyde in 0.1M Millonig’s buffer.
    Embedding : in Lowicryl 4KM at -20°C under UV.
    Ultrathin-sections deposited on formvar-coated carbonated EM-grids.
    Blocking step: 5% BSA for 30 seconds at RT.
    Primary antibody: ab1791 diluted 1/50 in PBS, for 1h at RT.
    Secondary antibody: BBI International anti-rabbit

Anti-Histone H3 antibody - Nuclear Loading Control and ChIP Grade (ab1791) 使用論文

This product has been referenced in:
  • Yang Q  et al. The Polycomb Group Protein EZH2 Impairs DNA Damage Repair Gene Expression in Human Uterine Fibroids. Biol Reprod N/A:N/A (2016). WB ; Human . Read more (PubMed: 26888970) »
  • Lima WF  et al. RNA cleavage products generated by antisense oligonucleotides and siRNAs are processed by the RNA surveillance machinery. Nucleic Acids Res 44:3351-63 (2016). Read more (PubMed: 26843429) »

See all 927 Publications for this product

Product Wall

Our antibody ab1791 will recognize total Histone H3 in your samples, so it will recognize the modified forms of Histone H3. Unfortunately I am unaware of the ratio of expression between the different variant forms in human cell lines.

I am happy to let you know that we have non-rabbit anti-human Histone H3 antibodies available, which also qualify for our testing discount programme (either because of their potential suitability for IF or because we do not have an image yet).
Read More

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Human Cell lysate - nuclear (B Cells)
Negative control IgG Pulldown
Specification B Cells
Detection step Real-time PCR
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: Formaldehyde
Positive control GAPDH Promoter
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Arunava Roy

Verified customer

投稿 Oct 27 2016

Application Western blot
Sample Monkey Cell lysate - whole cell (Kidney)
Gel Running Conditions Non-reduced Non-Denaturing (Native)
Loading amount 30 µg
Specification Kidney
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Oct 10 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - whole cell (Kidney)
Gel Running Conditions Non-reduced Non-Denaturing (Native) (4-12)
Loading amount 30 µg
Specification Kidney
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Oct 10 2016

Application Western blot
Sample Cow Cell lysate - whole cell (Lung)
Gel Running Conditions Non-reduced Non-Denaturing (Native)
Loading amount 30 µg
Specification Lung
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Oct 10 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Cell lysate - whole cell (neuroblastoma)
Gel Running Conditions Non-reduced Non-Denaturing (Native) (4-12)
Loading amount 30 µg
Specification neuroblastoma
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 4°C
Username

Abcam user community

Verified customer

投稿 Oct 10 2016

Application Western blot
Sample Human Cell lysate - whole cell (U2OS cells)
Gel Running Conditions Reduced Denaturing (4-12% Gradient Gel)
Loading amount 50 µg
Specification U2OS cells
Blocking step Licor Blocking Buffer as blocking agent for 30 minute(s) · Concentration: 100% · Temperature: 25°C
Username

Abcam user community

Verified customer

投稿 Oct 06 2016

Application ChIP
Sample Mouse Cell lysate - whole cell (gut)
Specification gut
Detection step Real-time PCR
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 10 minute(s) and 0 second(s)
Specification of the cross-linking agent: Formaldehyde
Username

Abcam user community

Verified customer

投稿 Sep 26 2016

Application Western blot
Sample Mouse Cell lysate - whole cell (primary cortical neurons)
Gel Running Conditions Reduced Denaturing (4-12% Bis-Tris)
Loading amount 10 µg
Specification primary cortical neurons
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

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投稿 Aug 17 2016

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