Human Histone H3 (tri methyl K4) peptide (ab1342)

Reviews (1)Q&A (5)References (18)
Dot Blot - Human Histone H3 (tri methyl K4) peptide (ab1342)

    Key features and details

    • Suitable for: Dot blot, Blocking

    製品の詳細

    特性

    Our Abpromise guarantee covers the use of ab1342 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

      Dot blot

      Blocking - Blocking peptide for Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580), Anti-Histone H3 (tri methyl K4) antibody [mAbcam12209] - ChIP Grade (ab12209)

    • 製品の状態

      Liquid

    • 備考

      - First try to dissolve a small amount of peptide in either water or buffer. The more charged residues on a peptide, the more soluble it is in aqueous solutions.
      - If the peptide doesn’t dissolve try an organic solvent e.g. DMSO, then dilute using water or buffer.
      - Consider that any solvent used must be compatible with your assay. If a peptide does not dissolve and you need to recover it, lyophilise to remove the solvent.
      - Gentle warming and sonication can effectively aid peptide solubilisation. If the solution is cloudy or has gelled the peptide may be in suspension rather than solubilised.
      - Peptides containing cysteine are easily oxidised, so should be prepared in solution just prior to use.

    • Concentration information loading...

    前処理および保存

    • 保存方法および安定性

      Shipped 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.

      Information available upon request.

    関連情報

    • 別名

      • H3 histone family member E pseudogene
      • H3 histone family, member A
      • H3/A
      • H31_HUMAN
      • H3F3
      • H3FA
      • Hist1h3a
      • HIST1H3B
      • HIST1H3C
      • HIST1H3D
      • HIST1H3E
      • HIST1H3F
      • HIST1H3G
      • HIST1H3H
      • HIST1H3I
      • HIST1H3J
      • HIST3H3
      • histone 1, H3a
      • Histone cluster 1, H3a
      • Histone H3 3 pseudogene
      • Histone H3.1
      • Histone H3/a
      • Histone H3/b
      • Histone H3/c
      • Histone H3/d
      • Histone H3/f
      • Histone H3/h
      • Histone H3/i
      • Histone H3/j
      • Histone H3/k
      • Histone H3/l
      see all

    • 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

    画像

    • Dot Blot - Human Histone H3 (tri methyl K4) peptide (ab1342)
      Dot Blot - Human Histone H3 (tri methyl K4) peptide (ab1342)This image is courtesy of an anonymous Abreview

      ab1342 (at 0.01 and 0.1 µg) used in Dot Blot to test for cross-reactivity of ab8580 (1/2500). An HRP-conjugated Mouse anti-rabbit polyclonal (1/10000) was used as the secondary antibody.

      See Abreview

    プロトコール

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    参考文献 (18)

    ab1342 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

    ab1342 は 18 報の論文で使用されています。

    • You Y  et al. Temporal dynamics of gene expression and histone marks at the Arabidopsis shoot meristem during flowering. Nat Commun 8:15120 (2017). PubMed: 28513600
    • Kinkley S  et al. reChIP-seq reveals widespread bivalency of H3K4me3 and H3K27me3 in CD4(+) memory T cells. Nat Commun 7:12514 (2016). PubMed: 27530917
    • McLaughlin N  et al. In situ histone landscape of nephrogenesis. Epigenetics 9:222-35 (2014). PubMed: 24169366
    • Rosselli JE  et al. The effect of simvastatin on the regeneration of surgical cavities in the femurs of rabbits. Acta Cir Bras 29:87-92 (2014). PubMed: 24604311
    • Huh YH & Sherley JL Decreased H3K27 and H3K4 trimethylation on mortal chromosomes in distributed stem cells. Cell Death Dis 5:e1554 (2014). PubMed: 25476902
    View all Publications for this product

    レビューと Q&A

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    1-6 of 6 Abreviews or Q&A

    Question

    I have checked up your products, however I cannot find the information at Abcam website about their residue sequences, residue positions and purity.
    I am doing research with the model plant Arabidopsis. I can use the antibodies and peptides derived from human, because histone H3 is mostly conserved among many eukaryotes, however there are still differences at some amino acid positions.

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    Answer

    The sequence and purification method is propriety, but if you let me know what is important for you I can check if one of our product fits.

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    Question

    Do you have any full size of H3K4 mono-, di- and tri methylated histone 3 which I can load on the gel as positive control.

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    Answer

    >The molecular weights of the different methylation states be extrememly difficult to distinguish as that type of modification creates a 14Dalton mass change.Currently, we do not offer full length methylated H3K4 proteins. We do have anEpiSeeker Histone Extraction Kit (ab113476) whichprovides a simple and selective method for extracting histone proteins in just 1 hour. When used in conjunction with our peptides you can be confident that the band you are seeing is indeed specific to the mono- di- or tri- methylayed H3K4 protein.

    More information about ourEpiSeeker Histone Extraction Kit (ab113476) is available here:

    https://www.abcam.com/EpiSeeker-Histone-Extraction-Kit-ab113476.html

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    Question

    We have your antibodies against Histone H3K4 methylation (ab8895 for monomethylation and ab8580 for trimethylation). I would like to see their specificity on WB. Do you have any WB data loading Histon H3 unmodified, monomethylated H3K4, dimethyated H3K4 and trimethylated H3K4 and then stained by ab8895 or ab8580 ? Also do you have any control peptide/protein applicable for WB of H3K4 methylation WB ? They should be full size of monomethylated H3K4, dimethylated H3K4, trimethylated H3K4 and unmodified H3.

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    Answer

    Peptides are available for both of these products. The peptide for ab8895,Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade, is product number ab1340 (https://www.abcam.com/histone-h3-peptide-mono-methyl-k4-ab1340.html).The peptide for ab8580,Anti-Histone H3 (tri methyl K4) antibody - ChIP Grade, is ab1342 (https://www.abcam.com/histone-h3-peptide-tri-methyl-k4-ab1342.html). These are the exact peptides used to create the antibodies. Each peptide used in the testing assay is available as well. (https://www.abcam.com/index.html?datasheet=1340,https://www.abcam.com/index.html?datasheet=7768),https://www.abcam.com/index.html?datasheet=1342,https://www.abcam.com/index.html?datasheet=1771,https://www.abcam.com/index.html?datasheet=1780,https://www.abcam.com/index.html?datasheet=2903.

    These products are tested by peptide assay. Our data on western blot is displayed on the datasheet for each product. For ab8895 our test show itspecific for mono-methylated Lysine 4 of histone H3 and does not recognize di- or tri-methyl Lysine 4 nor methylation at Lysine 9. In the image on the datasheet, this is shown in lane 2where the activity of the antibody is specifically blocked by the addition of the immunizing peptide (ab1340) [https://www.abcam.com/Histone-H3-mono-methyl-K4-antibody-ChIP-Grade-ab8895.html#ab8895_6.gif].

    Further,ab8580 are tested in Peptide Array against peptides to different Histone H3 modifications. Six dilutions of each peptide are printed on to the Peptide Array in triplicate and results are averaged before being plotted on to a graph. Results show strong binding to Histone H3 - tri methyl K4 peptide (ab1342), indicating that this antibody specifically recognises the Histone H3 - tri methyl K4 modification [https://www.abcam.com/Histone-H3-tri-methyl-K4-antibody-ChIP-Grade-ab8580.html#Histone-H3-Primary-antibodies-ab8580-153.jpg.]

    Each of these products are thoroughly characterized and published. There are links to specific publications and th Abreviews from our customers on each datasheet.

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    Other (Dot Blot) Abreview for Histone H3 peptide - tri methyl K4

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    Abreviews
    abreview image
    Application
    Other

    Review text: I used this product to do a dot blot specificity test for cross reactivity of modified histone antibodies and everything worked great. I made two dilutions of the peptide and spotted it on a nitrocellulose membrane using a Bio-Dot apparatus. Once the peptides were bound to the membrane I incubated with the appropriate primary and secondary antibodies at an assay dependent concentration and developed using ECL.
    Sample: Human Purified protein

    Primary antibody (in addition to 'Histone H3 peptide - tri methyl K4')
    Primary antibody: Abcam primary antibody: Anti-Histone H3 (methylated) (tri methyl K4) antibody (ab8580)
    Dilution: 1/2500

    Secondary antibody
    Name: Non-Abcam antibody was used: anti-rabbit
    Host species: Mouse
    Clonality: Polyclonal
    Conjugation: Horse Radish Peroxidase
    Dilution: 1/10000

    Abcam user community

    Verified customer

    投稿 Nov 01 2010

    Question

    I am writing to find out how the specificity of the antibodies I purchased were determined. I bought: Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895) Histone H3 (tri methyl K4) antibody - ChIP Grade (ab8580) Histone H3 (di methyl K4) antibody - ChIP Grade (ab7766) I also purchased these peptides: Histone H3 peptide (1-8) - tri methyl K4 (ab1342) Histone H3 peptide (1-10) - di methyl K4 (ab7768) Histone H3 peptide (1-8) - mono methyl K4 (ab1340) Histone H3 peptide (1-16) - unmodified (ab7228) I would like to know of any manual dot-blotting methods you have, that will work with these. I have tried to use them in a blot to no avail so far. The proteins did not bind to the PVDF membrane I used. I was attempting a procedure from the millipore company instructions. there is no dot-blotting apparatus in my lab, so I have to do it by hand.

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    Answer

    We do not have a recommended protocol for Dot-blotting but can recommend trying a nitrocellulose membrane, a general manual will be able to help you. Specificity of those antibodies was tested by WB of Histone (ab121) (0.5µg per lane) and with blocking peptides (modified and non modified peptides). In each case, blocking was achieved only when the adequate peptide was added.

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    Question

    You mention on your web site that your blocking peptides can be coupled to beads via their C-terminal Cys residu. Can you send me more info about that (what kind of beads, by what principle etc.)

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    Answer

    The mechanism for coupling the peptides to beads via their terminal cys residue is based on a maleimide reaction scheme. The maleimide group of crosslinkers such as sulfo SMCC (PIERCE) or iodoacetyl moieties on an immobilisation matrix such as Sulfolink (PIERCE) form a thioether bond with the free sulfhydyls in the cysteine residues of the peptides. The peptide can thus be immobilised on solid supports in this fashion and used for numerous applications.

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