Histone H3 (di-methyl K27) Quantification Kit (Colorimetric) (ab115070)

Submit a review Q&A (4)
Functional Studies - Histone H3 (di-methyl K27) Quantification Kit (Colorimetric) (ab115070)

    Key features and details

    • Assay type: Quantitative
    • Detection method: Colorimetric
    • Platform: Microplate reader
    • Assay time: 2 hr 30 min
    • Sample type: Adherent cells, Suspension cells, Tissue
    • Sensitivity: 2 ng/well

    Overview

    • Product name

      Histone H3 (di-methyl K27) Quantification Kit (Colorimetric)
      See all Histone H3 kits

    • Detection method

      Colorimetric

    • Sample type

      Tissue, Adherent cells, Suspension cells

    • Assay type

      Quantitative

    • Sensitivity

      2 ng/well

    • Range

      20 ng/well - 5000 ng/well

    • Assay time

      2h 30m

    • Species reactivity

      Reacts with: Mouse, Human
      Predicted to work with: Mammals

    • Product overview

      Methylation of histone H3 at lysine 27 (K27) is a mark of gene inactivation and it is thought to mediate heterochromatin formation by forming a binding site for Polycomb. H3 (methyl K27) is also thought to have an important role in the repression of HOX genes during development and in X chromosome inactivation and imprinting. H3 (di-methyl K27), a modification enriched at pericentromic heterochromatin, is observed to be broadly distributed throughout all euchromatic sites and participates in silencing gene expression.

      Histone H3 (di-methyl K27) Quantification Kit (Colorimetric) (ab115070) allows the user to specifically measure global di-methylation of histone H3K27 colorimetrically using a variety of mammalian cells including fresh and frozen tissues, cultured adherent and suspension cells.

    • Platform

      Microplate reader

    Properties

    • Storage instructions

      Please refer to protocols.
    • Components Identifier 48 tests 96 tests
      10X Wash Buffer 1 x 10ml 1 x 20ml
      8-Well Sample Strips (with Frame) 4 units 9 units
      8-Well Standard Control Strips Green Ringed 2 units 3 units
      Antibody Buffer 1 x 6ml 1 x 12ml
      Color Developer 1 x 5ml 1 x 10ml
      Detection Antibody, 1 mg/mL 1 x 5µl 1 x 10µl
      Standard Control, 100 µg/mL 1 x 10µl 1 x 20µl
      Stop Solution 1 x 3ml 1 x 6ml

    • Research areas

    • Function

      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.

    • Sequence similarities

      Belongs to the histone H3 family.

    • Developmental stage

      Expressed 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 localization

      Nucleus. Chromosome.
    • Information by UniProt

    • Alternative names

      • 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

    • Database links

      see all

    Images

    • Functional Studies - Histone H3 (di-methyl K27) Quantification Kit (Colorimetric) (ab115070)
      Functional Studies - Histone H3 (di-methyl K27) Quantification Kit (Colorimetric) (ab115070)

      ab115070 used to measure the amount of di-methyl H3K27. Histone extracts were prepared from MDA-MB-231 cells using the Histone Extraction Kit (ab113476).

    References (0)

    Publishing research using ab115070? Please let us know so that we can cite the reference in this datasheet.

    ab115070 has not yet been referenced specifically in any publications.

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-4 of 4 Abreviews or Q&A

    Question

    The credit was about ******* and this time, he has ******.
    These kits were not satisfied again, he already purchased other vendor's products and doesn't need to use ab115070, ab115072 any more.
    Please arrange credit note for our customer.

    Read More
    Answer

    Sorry for the confusion.

    As requested, I have asked our accounting department to issue you with a credit note. This can then be redeemed against the invoice of a future order.

    Credit ID: *****

    As usual if you have any further questions regarding this credit note, please contact the accounts department by email at creditcontrol@abcam.com. Please refer to the credit ID number in any correspondence with the accounting department.

    I would like to wish the customer good luck with their research.

    Read More

    Question

    Please arrange credit note for our customer.

    Read More
    Answer

    A credit note has already been issued on the DD MM YYYY for these two kits. The reference of the credit note is ********.

    In my previous email I said : "Would your customer agree to use their credit note to buy and try the kits for the 3rd time?"

    Read More

    Question

    As you know our customer is is havinga problem using you ab115070, ab115072.
    He recently received new kits and tested again, but he could not any results eventhoug at standard.
    So today I met him and checked his protocols, and there was no problem.
    When I saw this standar values (0-100ng), the OD value were almost same, it mean almost blank.
    So I tested in his lab, I used only blank , and 100ng standard. (Ihave many experiences)
    As a results I also could not see any differences in negative control and positive control.
    OD value were very low, there was no color change.
    I checked abcam's substate, so I mixed only non diluted detection antibody and substrate(1ul + 50ul), it worked.
    Next I checked diluted detection antibodies.
    When I mixed 1:1000 diluted antibody 10ul and substate, there was no color change.
    so I mixed 1:500 diluted antibody 10ul and substate, three was no color change.
    I tried other company's 2nd antibody, I diluted the antibody 1:1000 (HRP conjugated antibody) and add 5ul antibody and abcam's substrate.
    the color changed to blue, and when I added stop sol, the color changed to yellow.

    So I tested undiluted abcam's detection antibody (1ul) + 50ul substrate.
    there were little bit color change, but it is not strong, eventhouhg the antibody was non diluted.

    Could explain why these kits did not work?

    Read More
    Answer

    Please find below the last email I received from the lab where the kits are produced :

    "We are sorry to hear about it and not clear what caused it not work.
    The procedure seems fine.
    We did not receive any other complaint for the same lot of these 2 kits.
    Specifically, we tested the standard control for these two kits before sending them to you.
    In fact, the standard control and detection antibody are also used for other histone H3 modification assay kits as universal components.
    If the customer agrees, we couldsupply another kits whichare also specifically tested, again with good results."

    Read More

    Question

    As you know our customer is is havinga problem using you ab115070, ab115072.
    He recently received new kits and tested again, but he could not any results eventhoug at standard.
    So today I met him and checked his protocols, and there was no problem.
    When I saw this standar values (0-100ng), the OD value were almost same, it mean almost blank.
    So I tested in his lab, I used only blank , and 100ng standard. (Ihave many experiences)
    As a results I also could not see any differences in negative control and positive control.
    OD value were very low, there was no color change.
    I checked abcam's substate, so I mixed only non diluted detection antibody and substrate(1ul + 50ul), it worked.
    Next I checked diluted detection antibodies.
    When I mixed 1:1000 diluted antibody 10ul and substate, there was no color change.
    so I mixed 1:500 diluted antibody 10ul and substate, three was no color change.
    I tried other company's 2nd antibody, I diluted the antibody 1:1000 (HRP conjugated antibody) and add 5ul antibody and abcam's substrate.
    the color changed to blue, and when I added stop sol, the color changed to yellow.

    So I tested undiluted abcam's detection antibody (1ul) + 50ul substrate.
    there were little bit color change, but it is not strong, eventhouhg the antibody was non diluted.

    Finally I think abcam's ab115070, ab115072 have problem and this is not first time.
    Could explain why these kits did not work?
    I have to explant to our customer, he is working Korea NIH and very vaulable cusotmer to us.

    Read More
    Answer

    Thank you for contacting us.

    Thank you also for the detailed information about the use of the kits ab115070 and ab115072.

    I do believe there is a problem with those kits, so please send our sincere apologies to the customer.

    Once again I will pass on those details to the source of the kits, hoping the problem component(s) will be determined and corrected.

    I will contact you again as soon as I receive extra information forthe lab.

    Read More

    Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
    For licensing inquiries, please contact partnerships@abcam.com