Overview

  • Product name

    Anti-Histone H2A.Z antibody - ChIP Grade
    See all Histone H2A.Z primary antibodies
  • Description

    Rabbit polyclonal to Histone H2A.Z - ChIP Grade
  • Host species

    Rabbit
  • Tested applications

    Suitable for: ICC/IF, WB, IHC-P, ChIP, IPmore details
  • Species reactivity

    Reacts with: Human
    Predicted to work with: Schizosaccharomyces pombe
  • Immunogen

    Synthetic peptide corresponding to Human Histone H2A.Z aa 65-128.

  • Positive control

    • MOLT4 or Raji cell lysate. ICC/IF Hela cells IHC-P: Human normal colon FFPE tissue sections.

Properties

Applications

Our Abpromise guarantee covers the use of ab97966 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
ICC/IF Use a concentration of 0.5 µg/ml.
WB 1/500 - 1/3000. Predicted molecular weight: 14 kDa.
IHC-P Use a concentration of 0.1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
ChIP Use 2 µg for 25 µg of chromatin.
IP 1/500 - 1/1000.

Target

  • Function

    Variant histone H2A which replaces conventional H2A in a subset of nucleosomes. 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. May be involved in the formation of constitutive heterochromatin. May be required for chromosome segregation during cell division.
  • Sequence similarities

    Belongs to the histone H2A family.
  • Post-translational
    modifications

    Monoubiquitination of Lys-122 gives a specific tag for epigenetic transcriptional repression.
    Acetylated on Lys-5, Lys-8 and Lys-12 during interphase. Acetylation disappears at mitosis.
    Monomethylated on Lys-5 and Lys-8 by SETD6. SETD6 predominantly methylates Lys-8, lys-5 being a possible secondary site.
    Not phosphorylated.
  • Cellular localization

    Nucleus. Chromosome.
  • Information by UniProt
  • Database links

  • Alternative names

    • H2A histone family member Z antibody
    • H2A.z antibody
    • H2A/z antibody
    • H2afz antibody
    • H2AZ antibody
    • H2AZ_HUMAN antibody
    • Histone H2A.Z antibody
    • MGC117173 antibody
    see all

Images

  • Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab97966 (blue), and 20µl 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 for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

  • All lanes : Anti-Histone H2A.Z antibody - ChIP Grade (ab97966) at 1/3000 dilution

    Lane 1 : MOLT4 whole cell lysate
    Lane 2 : Raji whole cell lysate

    Lysates/proteins at 30 µg per lane.

    Predicted band size: 14 kDa



    12% SDS PAGE
  • IHC image of ab97966 staining Histone H3 (phospho T45) in human colon formalin fixed paraffin embedded tissue sections*, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab97966, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. No primary antibody was used in the negative control (shown on the inset).

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

    *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

  • ab97966 staining Histone H2A.Z 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 ab97966 at 0.5μg/ml overnight at +4°C, followed by a further incubation at room temperature for 1h with an AlexaFluor®488 Goat anti-Rabbit secondary (ab150077) at 2 μg/ml (shown in green). AlexaFluor®350 WGA was used at a 1/200 dilution and incubated for 1h with the cells, to label plasma membranes (shown in blue). Nuclear DNA was labelled in red with 1.25 μM DRAQ5™ (ab108410), which was added to the secondary antibody mixture. A secondary only negative control is displayed, which indicates that the Histone H2A.Z staining observed is due to primary antibody specificity and not to unspecific binding of the secondary antibody to the cells.

  • ab97966 immunoprecipitating Histone H2A.Z protein in HeLa whole cell lysate/extract. Lane 1: 50 μg HeLa whole cell lysate/extract. Lane 2: Control with 2 μg of preimmune rabbit IgG. Lane 3: Immunoprecipitation of Histone H2A.Z protein by 2 μg of ab97966. The immunoprecipitated Histone H2A.Z protein was detected with ab97966 diluted at 1:1000. Anti-rabbit IgG was used as a secondary antibody.

References

ab97966 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-7 of 7 Abreviews or Q&A

Application
ChIP
Sample
Schizosaccharomyces pombe Cell lysate - whole cell (Not applicable)
Negative control
No antibody added.
Specification
Not applicable
Detection step
Real-time PCR
Type
Cross-linking (X-ChIP)
Duration of cross-linking step: 15 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% Formaldehyde
Positive control
Used PCR primers cited in publications. These primers are specific for loci (Cox16 and 644.08c) that have been determined to be occupied by H2A.Z

Abcam user community

Verified customer

Submitted Sep 15 2018

Application
Western blot
Sample
Saccharomyces cerevisiae Cell lysate - whole cell (WT and htz1? (Htz1 deletion ) cells)
Gel Running Conditions
Reduced Denaturing (gel 15%)
Loading amount
1e+007 cells
Specification
WT and htz1? (Htz1 deletion ) cells
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 30°C

Daisuke Takahashi

Verified customer

Submitted Jul 17 2018

Question
Answer

Thank you very much for providing that information.

I am sorry that the antibody did not prove successful with your S. pombe samples. I do hope that you feel it was still worthwhile trying. As I mentioned in my previous email, please do go ahead and claim your free primary antibody (or the equivalent price of ab97966 off any product in our catalogue). As mentioned previously, along with the discount code could you also please quote "antibody unpublished, testing discount agreed with Karin" when placing your order. This means customer services know to speak to me once the order goes through so the discount will be added.

I hope this information has been of help. If I can be of any further assistance, please do let me know.

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Answer

I'm sorry about that. It might be that the system won't let you submit a review for an unpiblished antibody. I was not aware of this.

If you wouldn't mind, could you just share the information with me by filling in the relevant form attached to this email? This should only take 5 minutes. Could you also let me know how you would rate the antibody out of 5.

This information would only become accessible to others if we republish this antibody in the future.

Once the form has been returned to me, please go ahead and use the discount code I provided for you. Along with the discount code could you please quote "antibody unpublished, testing discount agreed with Karin". This means customer services know to speak to me once the order goes through so the discount will be added.

I look forward to receiving your reply.

Read More

Answer

Thank you for contacting me.

I am sorry that the Anti-Histone H2A.Z antibody (ab97966) has now been removed from the catalogue. I am sorry but I was not aware of this change when I suggested you test it. Let me reassure you that the antibody has not been removed due to any quality issues (if this were the case anyone who had purchased the affected antibody would have been informed).

You should still be able to submit an Abreview for the product. This can be done by following the link below:

https://www.abcam.com/index.html?pageconfig=resource&rid=10332&viapagetrap=abreviews

And clicking on the "https://www.abcam.com/index.html?pageconfig=your_reviews&strAccessMode=public_create".

You will need to log in to your account in order to achieve this. If you have any problems submitting the review please do let me know. As soon as the Abreview has been submitted, the discount code can be used to claim your free primary antibody (or the equivalent price of ab97966 off any product).

I hope this information has been of help. If you have any further questions please do let me know.

Read More

Answer

I am glad you found the information provided of help.

All you need to do in order to participate in the testing discount scheme is to purchase ab97966 as normal. Once you have used with yourS. pombe samples, please share the results observed by submitting an Abreview,regardless of whether they are positive or negative. In the Abreview form, under the "Additional comments" section please quote the following code:

Discount code: xxxxxxxx


Once the Abreview has been submitted, the code becomes active and you can then use it to claim a free primary antibody or the equivalent price of ab97966 off any product in our catalogue. This free product must be claimed within a 4 month period (by the 31st of October 2012).

More information on the testing discount scheme can be found from the following link:

https://www.abcam.com/collaboratordiscount

I hope I have explained the process sufficiently well for you. If you have any further questions, please do not hesitate to ask.

Otherwise, I wish you all the best with your experiments.

Read More

Answer

Thank you for contacting us and sorry for the delay in getting back to you.

I have checked the sequence provided and compared it to the immunogen of all the anti-Histone H2A.Z antibodies we have, including the ones you suggested. Unfortunately, the ones suggested by you all show very low homology between the immunogen used to raise the antibody and the S. pombe protein sequence. I would therefore not expect any of them to be able to detect the S. pombe protein.

I have however found one antibody that may be worth trying. This is the rabbit polyclonal ab97966. This has been raised againsta synthetic peptide taken from within the residues 65-128 of human Histone H2A.Z. The sequence alignment of this peptide (highlighted in yellow) can be viewed in the attachment of this email. As you can see the homology is very high, 93%. I would therefore expect the antibody to be able to detect the S. pombe protein. However, as we have not tested this ourselves I would not be able to guarantee it.

If you would like to purchase ab97966 for use with S. pombe samplesI would be able to offer you a testing discount.This offer means that if you purchase the antibody as normal and test the antibody for itsability to detect the S. pombe proteinby performing a Western blot, then let us know of the results through an Abreview (no matter whether positive or negative) you would be eligible for a free primary antibody of your choice from our catalog (or the equivalent price of ab97966 off any item in the catalogue). More information on this offer can be found here:

https://www.abcam.com/collaboratordiscount

If you would be interested in participating in this scheme please do let me know as a discount code needs to be issued prior to the purchase of ab97966.

I hope this information has been of help. If you require any further information please do not hesitate to contact us again.

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