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Rabbit specific HRP/DAB (ABC) Detection IHC Kit (ab64261)

Reviews (4)Q&A (13)References (107)
Immunohistochemistry - ab64261
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)

Overview

  • Product name

    Rabbit specific HRP/DAB (ABC) Detection IHC Kit
    See all HRP/DAB detection kit kits

  • Product overview

    Rabbit specific HRP/DAB (ABC) IHC Detection Kit ab64261 is a complete kit to detect a primary antibody raised in rabbit, using HRP-labeled-streptavidin and a biotinylated anti-rabbit secondary antibody.


    It is supplied with a full protocol and with peroxidase and protein blocking reagents, biotinylated secondary antibody, streptavidin-HRP, and DAB substrate.


    In the method:
    - a sample is incubated with an unconjugated primary antibody specific to the target antigen
    - a biotinylated secondary antibody then binds to the primary antibody
    - HRP-labeled streptavidin then binds to the secondary antibody
    - the HRP produces a brown colored substance at the site of primary antibody binding by reacting with DAB.

  • Notes

    For alternative streptavidin-biotin IHC detection kits see:
    - Anti-Mouse / anti-Rabbit HRP-DAB IHC kit ab64264
    - Anti-Mouse HRP-DAB IHC Kit ab64259

    For high sensitivity HRP micro-polymer IHC detection kits see:
    - Anti-Mouse / anti-Rabbit HRP/DAB Micro-polymer IHC Kit ab236466
    - Anti-Rabbit HRP-DAB Micro-polymer IHC Kit ab236469

    Alternatively, see HRP-AEC IHC detection kits.

    Find more kits and reagents for antigen retrieval, blocking, signal amplification, visualization, counterstaining, and mounting in the IHC kits and reagents guide.

  • Tested applications

    Suitable for: IHC-P, IHC-Frmore details

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 15 ml
    50x DAB Chromogen 1 x 0.5ml
    Biotinylated goat anti-rabbit IgG(H+L) 1 x 15ml
    DAB substrate 1 x 15ml
    Hydrogen Peroxide Block 1 x 15ml
    Protein block 1 x 15ml
    Streptavidin Peroxidase 1 x 15ml

  • Research areas

  • Relevance

    HRP/DAB detection kit is a immunoenzymatic antigen detection system for immunohistochemistry techniques.

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab64261 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
Use at an assay dependent concentration.
IHC-Fr
Use at an assay dependent concentration.
Notes
IHC-P
Use at an assay dependent concentration.
IHC-Fr
Use at an assay dependent concentration.

Images

  • Immunohistochemistry - ab64261
    Immunohistochemistry - ab64261Image from Narendra S.C., PLoS One 10(10), Fig 5a. doi: 10.1371/journal.pone.0141863 Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

    Immunohistochemical analysis staining TNF or IL6 in mouse tissue sections.  Samples are incubated in murine TNF (ab9739) or IL6 primary antibody. Bound primary antibodies were detected using IHC detection kit (ab64261). Immunohistochemistry was used to determine the effect of uridine on synovial expression of TNF and IL6. 

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)This image is courtesy of an anonymous Abreview

    Immunohistochemical analysis of KLF4 and pVHL expression in colon cancer tissues. Sections from colon cancer and adjacent tissues were analyzed by immunostaining against KLF4 and pVHL. Staining without primary antibody served as negative control.

    Tissue sections were de-waxed with xylene and rehydrated through gradient ethanol into water. For antigen retrieval, sections were heated in citrate buffer (pH6.0) for 10 min at 95°C in a microwave oven. After cooling to room temperature, the sections were then digested with 0.05% trypsin for 10 min at 37°C. Endogenous peroxidase activity was quenched with 0.3% H2O2 in methanol for 30 min at room temperature. After PBS washes, nonspecific antibody binding was blocked by pre-incubating slides with 10% normal goat non-immune serum at 37° for 30 min. After blotting off the blocking serum, sections were incubated with primary antibody against LKF4 as well as primary antibody against VHL at 4° overnight. After PBS washes again, sections were incubated with biotinylated secondary antibody at 1:200 dilution for 30 min at room temperature. After incubating with Vectastain ABC reagent (Vector Laboratories, Inc., Burlingame, CA) for 30 min at room temperature, the sections were developed with Anti Rabbit HRP/DAB Detection Kit (ab64261). Sections were washed in running tap water and lightly counterstained with hematoxylin, followed by dehydration and coverslip mounting. Negative controls were obtained by omitting the primary antibody.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti Rabbit HRP/DAB Detection Kit (ab64261)
    Human colon carcinoma fixed in 10% NBF for 24 hrs and stained with anti p53 antibody using Anti Rabbit HRP/DAB Detection Kit (ab64261).

References (107)

Publishing research using ab64261? Please let us know so that we can cite the reference in this datasheet.

ab64261 has been referenced in 107 publications.

  • Hou QQ  et al. TBC1D3 promotes neural progenitor proliferation by suppressing the histone methyltransferase G9a. Sci Adv 7:N/A (2021). PubMed: 33523893
  • Wu X  et al. DDB2 regulates DNA replication through PCNA-independent degradation of CDT2. Cell Biosci 11:34 (2021). PubMed: 33557942
  • Najdanovic JG  et al. Vascularization and osteogenesis in ectopically implanted bone tissue-engineered constructs with endothelial and osteogenic differentiated adipose-derived stem cells. World J Stem Cells 13:91-114 (2021). PubMed: 33584982
  • Hoshino K  et al. Interleukin-1ß Modulates Synaptic Transmission and Synaptic Plasticity During the Acute Phase of Sepsis in the Senescence-Accelerated Mouse Hippocampus. Front Aging Neurosci 13:637703 (2021). PubMed: 33643027
  • Shi W  et al. Reduction of TMAO level enhances the stability of carotid atherosclerotic plaque through promoting macrophage M2 polarization and efferocytosis. Biosci Rep 41:N/A (2021). PubMed: 33969376
View all Publications for this product

Customer reviews and Q&As

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11-17 of 17 Abreviews or Q&A

Research Associate

 Good Excellent 5/5 (Ease of Use)
Abreviews
abreview image
A review for the Rabbit specific HRP/DAB (ABC) Detection IHC Kit (ab64261)
Immunohistochemical staining procedure
Tissue sections were de-waxed with xylene and rehydrated through gradient ethanol into water. For antigen retrieval, sections were heated in citrate buffer (pH6.0) for 10 min at 95°C in a microwave oven. After cooling to room temperature, the sections were then digested with 0.05% trypsin for 10 min at 37°C. Endogenous peroxidase activity was quenched with 0.3% H2O2 in methanol for 30 min at room temperature. After PBS washes, nonspecific antibody binding was blocked by pre-incubating slides with 10% normal goat non-immune serum at 37° for 30 min. After blotting off the blocking serum, sections were incubated with primary antibody against LKF4 as well as primary antibody against VHL at 4° overnight. After PBS washes again, sections were incubated with biotinylated secondary antibody at 1:200 dilution for 30 min at room temperature. After incubating with Vectastain ABC reagent (Vector Laboratories, Inc., Burlingame, CA) for 30 min at room temperature, the sections were developed with DAB (Abcam, Cambridge, MA). Sections were washed in running tap water and lightly counterstained with hematoxylin, followed by dehydration and coverslip mounting. Negative controls were obtained by omitting the primary antibody.
Figure legend
Immunohistochemical analysis of KLF4 and pVHL expression in colon cancer tissues. Sections from colon cancer and adjacent tissues were analyzed by immunostaining against KLF4 and pVHL. Staining without primary antibody served as negative control.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Abcam user community

Verified customer

Submitted Jun 03 2014

Question

xxxx;

As I told you on the phone, I am planning on performing IHC on fresh PBMC samples from sheep.

Cells will be isolated from whole blood using a density gradient and will be placed on glass slides using a cytocentrifuge. At this point, we CAN fix them if needed, but we don’t have to. What do you recommend?

Our primary antibody is affinity purified rabbit polyclonal. We care considering using your EXPOSE Rabbit specific HRP-AES detection kit (ab94361) – do you think this is the best kit to use, or would you recommend a different one for our purposes? We are attempting to stain a transmembrane protein.

We are planning to counterstain using hematoxylin: do you think we would get the best contrast with the AEC kit or would you recommend using DAB?

Also, does abcam recommend purchasing hematoxylin from any particular vendor?

If ab64230 the appropriate mounting medium to use with this kit?

Thanks so much for your help. I look forward to your response!

Read More
Answer

Thank you for replying with that information.

As I mentioned over the phone, I have spoken to the source of these kits and we do not have any experimental data using either the ACE or DABdetection kits (ab64260, ab64261, ab80437 and ab94361)for IHC on fresh PBMC samples, asallof these kits have only been tested on formalin fixed paraffin embedded tissue samples.Wewould expect that as long as the primary antibody binds the target as expected that both kits could be suitable foryour samples but as we have not tested it ourselves we would not be able to guarentee this.



There should be good contrast if using the AEC or DAB kit with hematoxylin as counterstain. We counterstain with hematoxylin in-house regardless ofwhether DAB or AEC is used. The sensitivity betweenAEC and DABis similar. I would recommend using ab94361 or ab80437 if you expect the PBMC samples to contain any endogenous biotin.I am sorry but I cannot recommend a particular supplier of hematoxylin.

And to answer your final question, the mounting media ab64230 should be suitable for your purposes.

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

Read More

Question

Bonjour,

Tout d'abord merci pour votre aide.

J'ai effectué les tests avec votre kit de révélation pour la NGF et la TRkA avec les paramètres suivant :

- 3*5min a 600w dans un tampon à pH9

- 30 min de peroxydase

- 1h de serum block

- 30 min anticorps primaire

- 15 min biotine

- 15 min streptadivine

- DAB, Hematoxyline etc.

Nous avons tester sur le cerveau, sur la peau et sur de la muqueuse gingival. Nous avons un marquage mais nous ne sommes pas sur du résultat car dans la muqueuse l'épithélium est intégralement marqué. Est ce normal ?
De plus notre contrôle négatif, qui vient de chez vous,(ab47472) à aussi un léger font.

Les conditions si-dessus sont celle qui on donné le meilleur résultat. Pouvons nous vous envoyer les coupe afin que vous nous donniez votre avis ?

Merci d'avance pour votre aide.

Read More
Answer

Merci pour votre émail.

Je suis désolée que les résultats ne sont pas claires.

Je vais volontiers investiguer ces résultats et essayer de donner des conseils pour améliorer les images. Si un de nos produits serait défectueux, nous le remplacerons en accord avec notre garantie.

Pour mieux comprendre la problème, est-ce que vous pouvez s'il vous plaît m'envoyer les images annotées de ces résultats? C'est plus pratique si vous pouvez nous envoyer les images, et non les slides.

Aussi, pourriez vous s'il vous plait dire quels anticorps exactement vous avez utilisés pour les marquages avec quels kits? C'était le ab6199 ou le ab3125?

Est-ce que vous avez utilisé des coupes paraffine de souris? Quel sérum est-ce que vous avez utilisé pour le blocage? Est-ce que vous avez aussi utilisé le protéine block du kit?

Je me réjouis de votre réponse.

Read More

Question

Dear technical support Team, I am interested in the rabbit specific HRP/DAB detection IHC kit (ab64261). Does this kit work with ab6671 and ab6672 antibodies in mouse?

Read More
Answer

Thank you for contacting Abcam regarding our rabbit specific HRP/DAB detection IHC kit (ab64261) and our ab6671 and ab6672 antibodies. I have looked over the protocols and specifics of the kit and antibodies that you have mentioned. The antibodies have been tested for IHC of Formalin/PFA-fixed paraffin-embedded sections and shown to work in mouse tissues. I believe that you should encounter no difficulties in using ab6671 and ab6671 with this kit. All of our products come with the Abpromise guarantee. We will replace or refund products not performing as stated on the datasheet. Please do not hesitate to contact us with further questions or if there are other ways in which we might help you reach your research goals.      

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Question

Dears,   I want to know please, how many tests this kit can do? Thanks !   Kindest regards,    

Read More
Answer

Thank you for your enquiry. Where in the protocol it says 'cover the section', this usually means about 50 - 100 ul per slide.  Most of the reagents are provided at 15 ml, so this is enough for between 150-300 sections. I hope this will be helpful to you. If you have any further questions, please do not hesitate to contact us.  

Read More

Question

Can this staining kit be used for both frozen and paraffin-embedded sections?

Read More
Answer

ab64261 has not been validated for use on frozen sections, only for use on FFPE samples.  However, if the primary antibody has already been validated for use on frozens than I would expect that ab64261 would work on frozen sections.  You could probably start the protocol at step 4 for the frozen sections. I hope this is helpful. Please contact us again if you have any further questions.

Read More

Question

How many tests can you do with kit ab64261

Read More
Answer

Thank you for your recent enquiries regarding ab64261 Rabbit specific HRP/DAB detection IHC kit. I am sorry it has taken some time to confirm the details with our source. As discussed on the telephone, the number of tests that this kit can do is dependent on how much volume is used per slide. This will depends on the size of the tissue section(s). If you are using 100 ul/slide then the kit will do 150 tests. If you use 200 ul/slide it is good for 75 tests. For testing, our source uses 200 ul/slide. I hope this will be helpful to you. Should you have any further questions, please do not hesitate to contact us.

Read More

11-17 of 17 Abreviews or Q&A

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