Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)
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Overview
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Product nameAnti-Histone H3 (tri methyl K36) antibody - ChIP Grade
See all Histone H3 primary antibodies -
DescriptionRabbit polyclonal to Histone H3 (tri methyl K36) - ChIP Grade
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Host speciesRabbit
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SpecificityHistone H3 tri methylated at lysine 36
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Tested applicationsSuitable for: ICC/IF, CHIPseq, WB, ChIP, ChIP/Chip, IHC-Pmore details
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Species reactivityReacts with: Mouse, Rat, Cow, Human, Saccharomyces cerevisiae, Xenopus laevis, Arabidopsis thaliana, Caenorhabditis elegans, Drosophila melanogaster, Schizosaccharomyces pombe, Zebrafish, Silk worm, Rice, Xenopus tropicalis, Trypanosoma brucei
Predicted to work with: Plants
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Immunogen
Synthetic peptide within Human Histone H3 aa 1-100 (tri methyl K36) conjugated to Keyhole Limpet Haemocyanin (KLH). The exact sequence is proprietary.
(Peptide available asab1785) -
Positive control
- WB: Calf thymus histone preparation and HeLa whole cell extract. ICC/IF: HeLa cells.
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General notes
For detection of methylated histone H3.
Properties
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FormLiquid
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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.
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Storage bufferpH: 7.40
Preservative: 0.02% Sodium azide
Constituents: PBS, 1% BSA -
Concentration information loading... -
PurityImmunogen affinity purified
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Primary antibody notesFor detection of methylated histone H3
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ClonalityPolyclonal
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IsotypeIgG
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Research areas
Associated products
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ChIP Related Products
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Compatible Secondaries
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Control Peptide
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Immunizing Peptide (Blocking)
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Related Products
- Histone H3 (tri-methyl K36) Quantification Kit (Colorimetric) (ab115079)
- Prestained Protein Ladder - Broad molecular weight (10-245 kDa) (ab116028)
- Human Histone H3 (tri methyl K4) peptide (ab1342)
- Human Histone H3 (tri methyl K9) peptide (ab1773)
- Human Histone H3 (tri methyl K27) peptide (ab1782)
- Human Histone H3 (unmodified ) peptide (ab7228)
- Human Histone H3 (di methyl K4) peptide (ab7768)
Applications
Our Abpromise guarantee covers the use of ab9050 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 |
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| ICC/IF | Use a concentration of 0.1 - 1 µg/ml. We recommend using Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody. |
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| CHIPseq | Use at an assay dependent concentration. PubMed: 19581485 | |
| WB | Use a concentration of 1 µg/ml. Detects a band of approximately 15 kDa (predicted molecular weight: 15 kDa). | |
| ChIP | Use 4µg for 106 cells. | |
| ChIP/Chip | Use at an assay dependent concentration. | |
| IHC-P | Use a concentration of 0.5 - 10 µg/ml. |
Target
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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.
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Sequence similaritiesBelongs to the histone H3 family.
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Developmental stageExpressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
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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.
- Information by UniProt
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Database links
- Entrez Gene: 8290 Human
- 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
see all -
Alternative names
- HIST1 cluster, H3J antibody
- Histone gene cluster 1, H3 histone family, member E antibody
- Histone gene cluster 1, H3G antibody
see all
Images
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ChIP - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)Chromatin was prepared from U2OS 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 ab9050 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified on the GAPDH (active) and MYO-D (inactive) promoters and over the ý-Actin gene (active). Schematic diagram of the ý-Actin gene is shown on the top of the figure. Black boxes represent exons and thin lines represent introns. PCR products are depicted as bars under the gene.
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ChIP - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)This image is courtesy of an anonymous AbreviewChIP using ab9050 at the Pho4 locus in S. pombe. Chromatin extract from S.pombe cells was incubated with 5 ug of ab9050 overnight, and then incubated with Protein A beads for 1 hour. The immunoprecipitated DNA was quantified at the Pho4 locus by RT-PCR. No antibody was added to the beads control (red bars). A schematic diagram of the pho4 gene is shown below the graph. The grey box represents the open reading frame and PCR products are depicted by horizontal arrows. Please see anonymous abreview submitted on 29 October 2008 for additional details. -
Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)ab9050 stained in Hela cells. Cells were fixed with 100% methanol (5min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab9050 at 0.1µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilution overnight at +4°C. The secondary antibodies were Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) (pseudo-colored red) and Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed (ab150081) secondary antibody (colored green) used at 1 ug/ml for 1hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43µM for 1hour at room temperature.
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Western blot - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)All lanes : Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050) at 1 µg/ml
Lane 1 : Calf Thymus Histone Preparation Nuclear Lysate
Lane 2 : Calf Thymus Histone Preparation Nuclear Lysate with Histone H3 peptide - unmodified K36 at 0.5 µg/ml
Lane 3 : Calf Thymus Histone Preparation Nuclear Lysate withHuman Histone H3 (mono methyl K36) peptide (ab1783) at 0.5 µg/ml
Lane 4 : Calf Thymus Histone Preparation Nuclear Lysate withHuman Histone H3 (di methyl K36) peptide (ab1784) at 0.5 µg/ml
Lane 5 : Calf Thymus Histone Preparation Nuclear Lysate withHuman Histone H3 (tri methyl K36) peptide (ab1785) at 0.5 µg/ml
Lane 6 : Calf Thymus Histone Preparation Nuclear Lysate with Human Histone H3 (tri methyl K37) peptide (ab24417) at 0.5 µg/ml
Lysates/proteins at 0.5 µg per lane.
Secondary
All lanes : 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?) -
Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)ab9050 staining Histone H3 (tri-methyl K36) in HeLa cells. The cells were fixed with 100% methanol (5min) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab9050 at 0.1µg/ml and ab7291 (anti beta-Tubulin) at 1µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat anti -rabbit AlexaFluor®488 secondary (ab150081) at 2 μg/ml (shown in green) and a goat anti-mouse AlexaFluor®594 (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.
Negative controls: 1– Rabbit primary antibody and anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)This image is a courtesy of Anonymous Abreviewab9050 staining MAFA in Mouse Tissue sections (Liver, P5) by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde and blocked with 5% BSA for 1 hour at 25°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/100) for 3 hours at 25°C. An Alexa Fluor®555 conjugated Donkey anti-rabbit IgG polyclonal (1/250) was used as the secondary antibody.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)This image is courtesy of an anonymous Abreviewab9050 staining rat liver tissue sections by IHC-P. The section was formaldehyde fixed and subjected to heat mediated antigen retrieval in pH 6.0 citrate buffer prior to being blocked with 5% serum for 30 minutes at 45°C. The primary antibody was diluted 1/500 and incubated for 45 minutes at 20°C. A HRP conjugated goat anti-rabbit was used as the secondary
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Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (tri methyl K36) antibody - ChIP Grade (ab9050)This image is courtesy of an anonymous Abreviewab9050 staining Histone H3 (tri methyl K36) in Human Saos-2 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.25% Triton in PBS and blocked with 1% BSA for 1 hour at room temperature. Samples were incubated with primary antibody (1/1000) for 1 hour. An Alexa Fluor®488-conjugated Goat anti-rabbit IgG polyclonal (1/250) was used as the secondary antibody.
References
This product has been referenced in:
- Lu L et al. The plant-specific histone residue Phe41 is important for genome-wide H3.1 distribution. Nat Commun 9:630 (2018). Read more (PubMed: 29434220) »
- Moretto F et al. A regulatory circuit of two lncRNAs and a master regulator directs cell fate in yeast. Nat Commun 9:780 (2018). Read more (PubMed: 29472539) »
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