Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048)

Overview

  • Product nameAnti-Histone H3 (mono methyl K36) antibody - ChIP GradeSee all Histone H3 primary antibodies ...
  • Description
    Rabbit polyclonal to Histone H3 (mono methyl K36) - ChIP Grade
  • SpecificitySpecific for human Histone H3 mono methyl K36. Shows partial cross-reactivity with di-methyl K36 (please see Western Blot image). This antibody may not be suitable for experiments on yeast lysate. Although the antibody is specifically blocked using the immunising peptide, customer feedback indicates that it detects a band using S. cerevisiae K36 point mutants. We welcome further customer feedback.
  • Tested applicationsIHC-P, Flow Cyt, ICC, ICC/IF, WB, ChIP more details
  • Species reactivity
    Reacts with: Mouse, Cow, Human, Saccharomyces cerevisiae, Arabidopsis thaliana, Fruit fly (Drosophila melanogaster)
    Predicted to work with: all Mammals
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, mono methylated at K36.

    (Peptide available as ab1783.)

  • Positive control
    • HeLa Histone Preparation; Hela whole cell extract

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
  • Clonality Polyclonal
  • IsotypeIgG
  • Research Areas

Applications

Our Abpromise guarantee covers the use of ab9048 in the following tested applications.

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

Application Notes
IHC-P Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.
ICC Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration. PubMed: 19727073
WB 1/1000. Can be blocked with Human Histone H3 (mono methyl K36) peptide (ab1783).
ChIP Use a concentration of 1 - 4 µg/ml.

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.
  • Target information above from: UniProt accession P68431 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links
  • Alternative names
    • H3 histone family, member J antibody
    • FLJ92264 antibody
    • H3 histone antibody
    • H3 histone antibody
    • H3 histone family, member A antibody
    • H3 histone family, member B antibody
    • H3 histone family, member C antibody
    • H3 histone family, member D antibody
    • H3 histone family, member F antibody
    • H3 histone family, member H antibody
    • H3 histone family, member I antibody
    • H3 histone family, member K antibody
    • H3 histone family, member L antibody
    • H3 histone, family 3A antibody
    • H3.3A antibody
    • H3/a antibody
    • H3/b antibody
    • H3/c antibody
    • H3/d antibody
    • h3/f antibody
    • H3/h antibody
    • H3/i antibody
    • H3/j antibody
    • H3/k antibody
    • H3/l antibody
    • H31_HUMAN antibody
    • H3F1K antibody
    • H3F3 antibody
    • H3F3 antibody
    • H3FA antibody
    • H3FB antibody
    • H3FC antibody
    • H3FD antibody
    • H3FF antibody
    • H3FH antibody
    • H3FI antibody
    • H3FJ antibody
    • H3FK antibody
    • H3FL antibody
    • HIST1H3A antibody
    • HIST1H3B antibody
    • HIST1H3C antibody
    • HIST1H3D antibody
    • HIST1H3E antibody
    • HIST1H3F antibody
    • HIST1H3G antibody
    • HIST1H3H antibody
    • HIST1H3I antibody
    • HIST1H3J antibody
    • HIST3H3 antibody
    • HIST3H3 antibody
    • Histone 1, H3a antibody
    • Histone 1, H3b antibody
    • Histone 1, H3c antibody
    • Histone 1, H3d antibody
    • Histone 1, H3e antibody
    • Histone 1, H3f antibody
    • Histone 1, H3g antibody
    • Histone 1, H3h antibody
    • Histone 1, H3i antibody
    • Histone 1, H3j antibody
    • Histone cluster 1, H3a antibody
    • Histone cluster 1, H3b antibody
    • Histone cluster 1, H3c antibody
    • Histone cluster 1, H3d antibody
    • Histone cluster 1, H3e antibody
    • Histone cluster 1, H3f antibody
    • Histone cluster 1, H3g antibody
    • Histone cluster 1, H3i antibody
    • Histone cluster 1, H3j antibody
    • Histone H 3 antibody
    • Histone H3.1 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 (mono methyl K36) antibody - ChIP Grade images

  • ab9048 staining Histone H3 (mono methyl K36) in human differentiated haematopoietic stem cells by Flow Cytometry. Cells were fixed with paraformaldehyde and permeabilized with permeabilization buffer. The sample was incubated with the primary antibody (1/500) for 12 hours at 4°C. An Alexa Fluor® 488-conjugated goat polyclonal anti-rabbit IgG (1/1000) was used as the secondary antibody.
    Gating Strategy: Isotype negative control (white).

    See Abreview

  • All lanes : Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048) at 1/500 dilution

    Lane 1 : Histone prep
    Lane 2 : Histone prep with Human Histone H3 (mono methyl K36) peptide (ab1783) at 1 µg/ml
    Lane 3 : Histone prep with Human Histone H3 (di methyl K36) peptide (ab1784) at 1 µg/ml
    Lane 4 : Histone prep with Human Histone H3 (tri methyl K36) peptide (ab1785) at 1 µg/ml
    Lane 5 : Histone prep with Human Histone H3 (unmodified ) peptide (ab2623) at 1 µg/ml
    Lane 6 : Histone prep with Human Histone H3 (mono methyl K4) peptide (ab1340) at 1 µg/ml

    Lysates/proteins at 0.5 µg per lane.

    Secondary
    Goat Anti-Rabbit IgG H&L (HRP) (ab6721) at 1/5000 dilution

    Secondary ab: Alexa Fluor 680 Goat anti-rabbit IgG

    0.5µg histone prep used per lane

    Primary antibody:
    Lane 1: ab9048 (Histone H3 Mono Methyl K36) 1/500
    Lane 2: ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1783 (ab9048) (Histone H3 Mono Methyl K36) peptide 1µg/ml
    Lane 3: ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1794 (ab9049) (Histone H3 Di Methyl K36) peptide 1µg/ml
    Lane 4: ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1785 (ab9050) (Histone H3 Tri Methyl K36) peptide 1µg/ml
    Lane 5: ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab2623 (Histone H3 (23-34) – unmodified) peptide 1µg/ml
    Lane 6: ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1340 (ab8895) (Histone H3 Mono methyl K4) peptide 1µg/ml

    ab9048 specifically recognise

  • Staining of interphase nuclei of Hela cells with ab9048 (green) at a working dilution of 1/500. The DNA is stained with DAPI. ab9048 appears to be more associated with heterochromatin (DAPI intense regions) than euchromatin (DAPI less intense regions).

  • 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  ab9048 (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.

  • ab9048 staining Histone H3 (mono methyl K36) in Mouse pancreatic tumor tissue by Immunohistochemistry (Formalin/ PFA-fixed paraffin-embedded tissue sections). The sections were PFA-fixed and subjected to Heat mediated antigen retrieval in citrate and permeabilized with PBST prior to blocking with 5% serum for 1 hour at room temperature. The primary antibody was diluted 1/500 in PBS and incubated with the sample for 8 hours at 4°C. A Biotin-conjugated Goat anti-Rabbit polyclonal was used as the secondary antibody, diluted 1/1000.

    See Abreview

  • ICC/IF image of ab9048 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab9048, 0.1µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% formaldehyde fixed (10 min) Hek293, HepG2 and MCF7 cells at 0.1µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 0.1µg/ml.
  • All lanes : Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048) at 1/1400 dilution

    Lane 1 : Whole cell lysate prepared from Drosophila BG3 cells
    Lane 2 : Whole cell lysate prepared from Drosophila BG3 cells

    Lysates/proteins at 500000 cells per lane.

    Secondary
    HRP donkey anti-rabbit monoclonal at 1/20000 dilution
    developed using the ECL technique

    Observed band size : 17,42 kDa (why is the actual band size different from the predicted?)


    Exposure time : 30 seconds

    Image courtesy of an anonymous Abreview.

    See Abreview

References for Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048)

This product has been referenced in:
  • van Nuland R  et al. Nucleosomal DNA binding drives the recognition of H3K36-methylated nucleosomes by the PSIP1-PWWP domain. Epigenetics Chromatin 6:12 (2013). Read more (PubMed: 23656834) »
  • Jaffe JD  et al. Global chromatin profiling reveals NSD2 mutations in pediatric acute lymphoblastic leukemia. Nat Genet 45:1386-91 (2013). Read more (PubMed: 24076604) »

See all 27 Publications for this product

Product Wall

Application Flow Cytometry
Fixation Paraformaldehyde
Permeabilization Yes - eBioscience Permeabilization Buffer
Sample Human Cell (Differentiated Haematopoietic Stem Cells)
Specification Differentiated Haematopoietic Stem Cells
Gating Strategy Isotype negative control (white)
Preparation Cell harvesting/tissue preparation method: Typsin-EDTA Cell Dissociation
Sample buffer: PBS and 10% FBS
Username

Abcam user community

Verified customer

Submitted Feb 20 2014

Application Western blot
Loading amount 20 µg
Gel Running Conditions Reduced Denaturing
Sample Human Tissue lysate - whole (Patient Dermal Fibroblasts)
Specification Patient Dermal Fibroblasts
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Feb 01 2014

Application Western blot
Loading amount 4.5 µg
Gel Running Conditions Reduced Denaturing (12)
Sample Human Cell lysate - nuclear (Hela cells)
Specification Hela cells
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Oct 14 2013

Thank you for contacting us.
I apologize for the delays as I thought I have sent you an answer. I have had discussion with laboratory for the smaller sizes of ab9050 and ab9049. Actually they won't be able to treat these abs as a panel and wouldn’...

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Thank you for contacting us.
We are considering your request as a priority. Could you answer the following questions;
1. How many micro litres of antibody are required?
2. would this be 1 unit of each ab supplied at a smaller size?
3. ...

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Merci de nous avoir contactés.

J’ai vérifié les produits mentionnés : ab9048, ab9050 et ab89906. Puisque ces produits ont une concentration >1mg/ml de BSA, je peux vous confirmer qu’ils ne contienne...

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Merci de nous avoir contactés.

L’anticorps ab9048 est vendu dans du tampon sans BSA et il est en stock.

Nous prenons nos plaintes très au sérieux et donc pour les autres anticorps mentionné, pouvez-vo...

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Thank you for contacting us and your interest in our products.


Our anti-histone antibodiesare generally blocked with 5% BSA in 0.1% TBST for 1 hour at room temperature with gentle rocking when performing western blotting. We also use 5%...

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Thanks for your reply, and I apologize again for the delay getting this data to you.

Please see attached for the data for both lots, and if you have any questions or concerns please contact me and I'll be happy to help.

Have a great...

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Sure, I will get in touch with the lab and request that information for you. I apologize for the delay, but I expect to have all of the data by tomorrow.

Please let me know if you need anything else in the meantime and I'll be happy to help.

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