Overview

  • Product name
    Anti-8 Hydroxyguanosine antibody
    See all 8 Hydroxyguanosine primary antibodies
  • Description
    Goat polyclonal to 8 Hydroxyguanosine
  • Host species
    Goat
  • Specificity
    ab10802 cross reacts completely with 8-OHdG,and does not cross react with other naturally occurring nucleotides.
  • Tested applications
    Suitable for: IHC-P, ELISAmore details
  • Species reactivity
    Due to the conserved nature of the target reactivity would be predicted in wide range of species.
  • Immunogen

    8-Hydroxyguanosine - conjugate.

  • Positive control
    • Stressed tissue such as Alzheimer's affected neurons. or Human brain pre treated with 10ug proteinase K for 40mins at 37oC

Properties

  • Form
    Liquid
  • Storage instructions
    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.
  • Storage buffer
    Preservative: 0.09% Sodium azide
    Constituent: Whole serum
  • Purity
    Whole antiserum
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab10802 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
IHC-P 1/200.

Perform enzymatic antigen retrieval with 10 µg/ml proteinase K for 40 minutes at 37°C.

ELISA 1/100000 - 1/250000.

Target

  • Relevance
    8-Hydroxydeoxyguanosine (8OHdG) is a modified base that occurs in DNA due to attack by hydroxyl radicals that are formed as byproducts and intermediates of aerobic metabolism and during oxidative stress. There is increasing evidence to support the involvement of free radical reactions in the damage of biomolecules that eventually lead to several diseases in humans, such as atherosclerosis, cerebral and heart ischemia-reperfusion injury, cancer, rheumatoid arthritis, inflammation, diabetes, aging, and neurodegenerative conditions, such as Alzheimer’s disease. 8OHdG has become increasing popular as a sensitive, stable and integral marker of oxidative damage in cellular DNA. Biomonitoring in humans has demonstrated that 8OHdG can be excreted in the urine, and that a significant increase is caused by exposure to tobacco smoke and ionizing radiation. Because 8OHdG is so well correlated with oxidative stress and damage to DNA, which leads to degenerative disease states, the development of an antibody that can be used to study DNA damage has numerous applications. In addition to the direct study of DNA damage within cells, this antibody has applications in the development of immunoassays that can monitor 8OHdG excretion in the urine and serve as a biomarker of oxidative stress.
  • Alternative names
    • 8 hydroxyguanine antibody
    • 8 hydroxyguanosine antibody
    • 8 OHG antibody
    • 8-OHG antibody
    • 8OG antibody
    see all

Images

  • Immunohistochemical analysis of cat hepatic tissue, labeling 8 Hydroxyguanosine with ab10802. Sample fixed in paraformaldehyde. Heat mediated antigen retrieval with Tris-EDTA (pH9). Treated with ab10802 diluted 1/1500 in PBS plus casein for 1 hour and 30 minutes at 37°C. Secondary was ImmPress anti-goat HRP-conjugated polyclonal antibody.

    See Abreview

  • IHC staining on Alzheimer disease brain showing oxidized RNA in neurons. The tissue sections were formalin-fixed, paraffin embedded with either (A) no pretreatment or (B) pretreatment with 10 µg/ml proteinase K for 40 minutes at 37°C. Each was stained with a 1:200 dilution.

References

This product has been referenced in:
  • Yu H  et al. Mitochondrial Molecular Abnormalities Revealed by Proteomic Analysis of Hippocampal Organelles of Mice Triple Transgenic for Alzheimer Disease. Front Mol Neurosci 11:74 (2018). Read more (PubMed: 29593495) »
  • Wang D  et al. TRPC1 Deletion Causes Striatal Neuronal Cell Apoptosis and Proteomic Alterations in Mice. Front Aging Neurosci 10:72 (2018). Read more (PubMed: 29615894) »
See all 26 Publications for this product

Customer reviews and Q&As

1-10 of 22 Abreviews or Q&A

Answer

Thank you for contacting us.

I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

In order to better understand the problem, I’d like to send you a questionnaire with some questions about the protocol used.

Once we receive the completed questionnaire, we will look at the protocol and see if there are any suggestions we can make that may improve the results.

I would also appreciate if you could send us an image any of the failed experiments, to better understand the problem.

I look forward to receiving your reply. Please do not hesitate to contact us if you need any more advice or information.

Read More
Abcam has not validated the combination of species/application used in this Abreview.
Application
IHC - Wholemount
Sample
Mouse Tissue (colon)
Specification
colon

Dr. K Kulmira Nurgali

Verified customer

Submitted Nov 10 2017

Answer

This product has not been purified and is sold as a whole antiserum therefore we have not determined its concentration. As a rough guideline, in general whole antiserum has a concentration of 1-10 mg/ml.

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Answer

The antibody will detect 8-OHdG in DNA and also RNA. RNase digestion recommended if you want to detect only 8-OHdG in DNA.

The paper at the following link used a different antibody, a mouse monoclonal antibody, but the procedure for staining and digesting with RNase will be the same. You may want to include a DNase control, as they do in the paper, to demonstrate staining of DNA versus RNA.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1866598/

Am J Pathol. 1999 May;154(5):1423-9. PMID: 10329595

The staining procedure for ab10802 includes a pre-digestion with proteinase K for antigen retrieval, 10ug/ml for 40 minutes at 37C. You may need to test different incubation times (for instance, 0, 10, 20, and 40 minutes) to see which gives the best result.

Alternatively, the procedure used in the paper at the following link used a heat-mediated antigen retrieval step, instead of enzymatic retrieval, with EDTA buffer. They did not give a recipe for the EDTA buffer but it is typically 1mM EDTA, 0.05% Tween 20, pH 8.0, and the treatment consists of heating the sections in this buffer at 95-100C for 20 minutes, and then allowing the buffer to cool for 20 minutes before continuing with the IHC.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3122372/

Brain. 2011 Jul;134(Pt 7):1914-24. PMID: 21653539

Here is a link to a standard protocol for EDTA antigen retrieval.

http://www.ihcworld.com/_protocols/epitope_retrieval/edta.htm

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Answer

The immunostain that used the antibody at 1/4000 used a biotin-conjugated secondary, so it is likely that this was followed with an avidin-biotin-HRP complex amplification. It is possible that 1/4000 was in fact effective, as antibody affinities vary a great deal.

Another example on the datasheet, the immunofluorescent stain of mouse brain, used the antibody at 1/1000. (PubMed: 22776356 and PubMed: 21824519) So, the optimal dilution will depend on your sample and what you use for a secondary detection.

I suggest, if you are using an anti-mouse IgG or IgG2a -specific secondary directly conjugated to HRP, that you try 1/250, 1/500, and 1/1000. If you want to try just one dilution, then try 1/500 but closely monitor development of the substrate over time for just one slide before adding substrate to the other sections, to limit potential over-staining and background.

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Answer

Are you confident the secondary antibody and other reagents are effective? If so, I agree that it is best to try a different antibody at this point, which I will send free-of-charge.

Abcam has several others against 8-Hydroxyguanosine but as you may have noticed, the ones that have the most data and references are mouse monoclonals. To detect these will require an anti-mouse IgG secondary, which is likely to react with endogenous mouse IgG in the lung. If you already have an anti-mouse IgG secondary, you could test this to see if it will in fact be a problem.

One approach for minimizing the signal contributed by an anti-mouse secondary antibody is to choose one that is specific for the isotype sub-class of the primary, and that will not react with other IgG sub-classes. For example, if you choose anti-8 Hydroxyguanosine antibody ab62623, which is isotype IgG2a,

https://www.abcam.com/index.html?datasheet=62623

then you would pair it with an anti-mouse IgG2a that has been purified to remove reactivity with other subclasses such as IgG1. An example of this is ab98698:

https://www.abcam.com/index.html?datasheet=98698

This secondary antibody is conjugated to HRP, but we have other conjugates if you need, for instance, a fluorescent signal.

Alternatively, I can send another goat polyclonal such as ab93701:

https://www.abcam.com/index.html?datasheet=93701

or a refund.

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Answer

Thank you for your enquiry.

I can confirm that ab10802 Anti-8 Hydroxyguanosine antibody is tested as suitable for indirect ELISA.

However, I am sorry to confirm that as far as we are aware, ab10802 has never been tested in flow cytometry. All tested applications covered by the 6 month guarantee specified on our datasheets, and these are updated as soon as any new information is brought to our attention.

If you would like to test the antibody in flow cytometry, please do not hesitate to contact me again prior to the purchase by replying to this message as you may be eligible for our testing discount program.

Otherwise, we like to encourage all of our customers to submit an Abreview via the online product datasheet. We always appreciate customer feedback, whether positive or negative, and we make all product information available to researchers. Plus, each Abreview earns Abpoints that can be used for discounts on future purchases or rewards such as Amazon.com gift certificates.

To find out more about our Abreview system, please see the following link:

https://www.abcam.com/abreviews

Regarding the secondary antibodeis, I can recommend the following will be suitable for use with ab10802. These are anti Goat IgG secondary antibodies for detection of goat IgG primary.


For fluorescent ELISA or flow cytometry:
ab7121 Donkey anti-Goat IgG H&L (FITC) secondary antibody
FITC conjugated
Tested in: Flow Cyt, IHC-P, ELISA, IM, ICC/IF, IHC-Fr
https://www.abcam.com/index.html?datasheet=7121 (or use the following: https://www.abcam.com/index.html?datasheet=7121).
Enzymatic detection for ELISA:

ab97120 Donkey anti-Goat IgG H&L (HRP) secondary antibody
HRP conjugated
Tested in: IHC-P, ICC, WB, ELISA
https://www.abcam.com/index.html?datasheet=97120 (or use the following: https://www.abcam.com/index.html?datasheet=97120).

We have further information regarding applicatinos and protocols on the follwoing page from our website which you may find useful:
https://www.abcam.com/index.html?pageconfig=popular_protocols


I hope this information is helpful. Please do not hesitate to contact me for any further advice or information.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Cat Tissue sections (liver)
Specification
liver
Fixative
Paraformaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Tris-EDTA (pH 9.0)
Permeabilization
No
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 2.5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Feb 04 2013

Answer

Thank you for contacting us.

My Spanish speaking colleague is currently out of the office, unfortunately my Spanish is not good enough to write my reply in your language.

As stated on the product datasheets, both anti-8 Hydroxyguanosine antibodies ab26842 and ab10802 are guaranteed to work in IHC-P and ICC/IF.

The taget 8 Hydroxyguanosine is not different from one species to another like a protein would be. This is why there is no list of tested species for those antibodies.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

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Answer

I appreciate the time taken to submit further information to us.

Often it is possible to make suggestions that help resolve problems. We will happily offer technical support and in the event that a product is not functioning in the applications cited on the product data sheet (and the problem has been reported within 6 months of purchase), and if it appears that the antibody is at fault, a replacement/credit note/refund will be offered.

High background could be due to numerous reasons. In order to optimise the protocol and obtain better results from this antibody I would suggest the following steps:

- Blocking conditions. The secondary antibody may cross react with endogenous immunoglobulins in the tissue. This is avoided by pre-treating the tissue with normal serum from the species in which the secondary was raised. 1-2 hours of tissue blocking at RT could eliminate most of the background.

- Has endogenous peroxidase been blocked? In case it hasn’t it is also highly recommended to avoid cross reactions.

- As the expected staining is predicted to be nuclear, it may be worth trying a permeabilization step to allow the antibody reach the epitope. Triton or NP-40 0.1 to 0.2% for 10min are recommended agents for dissolving the nuclear membrane.

- Running a no primary control is usually very helpful to assess the source of the background.

I hope this information is helpful. Should the suggestions not improve the results, please do not hesitate to contact me again and I will try to provide further help.

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1-10 of 22 Abreviews or Q&A

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