製品の概要

  • 製品名
  • サンプルの種類
    Adherent cells, Suspension cells
  • 製品の概要

    ChIP kit ab500 provides a protocol and reagents for running ChIP assays including:
    - cell lysis and chromatin extraction
    - chromatin shearing and DNA fragment length analysis
    - immunoprecipitation and DNA purification


    DNA produced using the kit can be analyzed using qPCR or sequencing.


    The kit has been validated for ChIP assays with mammalian samples.

  • 特記事項

    This kit uses Protein A sepharose beads for antibody pulldown. See table below for Protein A and Protein G binding affinities with antibodies from commonly used species. For other species of antibody, consult the table in the protocol booklet.

    Species raised in

    Isotype  Protein A binding affinity Protein G binding affinity
           
    Rabbit  All isotypes +++ ++
           
    Goat All isotypes - ++
           

     

     

    Mouse   

    IgG1  + +++ 
    IgG2a  +++ +++
    IgG2b  ++ ++ 
    IgG3  +  +
    IgM  Use anti-mouse IgM 
  • アプリケーション
    適用あり: ChIPmore details

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab500 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
ChIP Use at an assay dependent concentration.

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  • Chromatin immunoprecipitation assay was performed to define the interaction of serum response factor (SRF) with the intragenic serum response element (SRE) regulatory motif in mouse using ab500 ChIP Kit

  • Chromatin immunoprecipitation using ab500 ChIP Kit and Histone H3 (tri methyl K9) antibody (ab8898). Chromatin was prepared from HeLa cells using the Abcam ChIP kit protocol. Cells were fixed with formaldehyde for 10 min. ChIP was performed with 2 µg of ab8898 (blue). No antibody was added to the beads control (yellow). Immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active/inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of transcribed region.
  • Chromatin immunoprecipitation using ab500 ChIP Kit and Histone H3 (tri methyl K4) antibody (ab12209). Chromatin was prepared from HeLa cells using the Abcam ChIP kit protocol. Cells were fixed with formaldehyde for 10 min. ChIP was performed with 5 µg of ab12209 (blue). No antibody was added to the beads control (yellow). Immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active/inactive loci). Primers and probes are located in the first kb of the transcribed region.
  • Chromatin immunoprecipitation using ab500 ChIP Kit and Histone H3 antibody (ab1791). Chromatin was prepared from HeLa cells using the Abcam ChIP kit protocol. Cells were fixed with formaldehyde for 10 min. ChIP was performed with 2 µg of ab1791 (blue). No antibody was added to the beads control (yellow). Immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active/inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

プロトコール

参考文献

This product has been referenced in:
  • Lorentzen A & Mitchelmore C NDRG2 gene copy number is not altered in colorectal carcinoma. World J Clin Oncol 8:67-74 (2017). ChIP ; Human . Read more (PubMed: 28246586) »
  • Albert BJ  et al. Combinations of isoform-targeted histone deacetylase inhibitors and bryostatin analogues display remarkable potency to activate latent HIV without global T-cell activation. Sci Rep 7:7456 (2017). Read more (PubMed: 28785069) »

See all 36 Publications for this product

レビューと Q&A

DNA purification slurry is often used for DNA extraction. The extraction is set up under aqueous alkaline conditions. In this environment, the slurry has an increased affinity for heavy metal cations such as Ca2+, Mn2+, and Mg2+. Nuclear DNA and RNA re...

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Thank you for your reply.

I will do my best to see if I can help you:

1 - When you analyze your results via PCR rather than western blotting, do you get the correct results (eg no signal in the negative control)? If you have not don...

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The use of ab500 ChIP kit has not been validated so far on bacteria.

The part of the protocol that may need some optimization is the chromatin preparation. Actually, lysis of bacteria is more complex than lysis of eukaryotic cells and may...

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the concentration of formaldehyde needed for the kit is actually 1% but you could use any percentage below 4%. 1% is just what was used in our lab.

It is possible to freeze the sheared chromatin and resume the ChIP at a later time, but once the ChIP has started, it is not possible to stop.

The kit is for 24 assays. It is 100 µl per ChIP. With 3000 µl DNA purifying slurry, you can do 30 ChIPs. But you are correct, you have to also use 100 µl per INPUT. So with this kit, you can do 24 assays + 6 INPUT.

Thank you for your inquiry.

I am happy to confirm that it is normal to have precipitation at 4°C in buffer D. This is due to the SDS as you said in your email.

It is important to bring the buffer to room temperature before addin...

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Thank you for your enquiry regarding ab500.

We never tested this kit on FUNGI chromatin. You may need to optimize first the shearing conditions. Afterwards, if you use a specific antibody to FUNGI chromatin, the kit will probably work.
<...

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Buffer D contains a high concentration of SDS. So, sometimes with low temperature, crystals will begin to form in this buffer. We recommend making sure that there are no crystals in the Buffer D by gently heating and mixing until crystals disappear.

Thank you for contacting abcam.

Earlier we spoke about your use of ab500 ChIP kit in conjunction with ab10812. You mentioned that the kit appears to work without the antibody added, but when using both ab10812 and the beads you do not get any ...

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1-10 of 46 Q&A

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

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