Overview

  • Product name
    DAB Substrate Kit
  • Product overview

    DAB substrate kit for IHC staining using peroxidase-based detection (ab64238). Ideal for autostainers as reagent is stable for up to 6 hours after mixing the two components.


    3,3'Diaminobenzidine (DAB) is a widely used chromogen for immunohistochemical staining. In the presence of peroxidase enzyme, DAB produces a brown precipitate that is insoluble in alcohol.


    This product is a two component form consisting of a liquid, refrigerator stable DAB Chromogen and DAB Substrate. 

  • Notes

    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-Pmore details

Properties

  • Storage instructions
    Store at +4°C. Please refer to protocols.
  • Storage buffer
    Constituents: 100% Propylene glycol, 5% Hydrogen peroxide, DAB substrate buffer
  • Components 60 ml 125 ml
    50x DAB Chromogen 1 x 2ml 1 x 4ml
    DAB substrate 1 x 60ml 1 x 125ml
  • Research areas
  • Relevance
    3,3' Diaminobenzidine (DAB) is a widely used chromogen for immunohistochemical staining. In the presence of peroxidase enzyme, DAB produces a brown precipitate that is insoluble in alcohol.
  • Alternative names
    • 3 3' Diaminobenzidine

Applications

Our Abpromise guarantee covers the use of ab64238 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 dilution.

Images

  • Immunohistochemical analysis of healthy and colitis mouse colon sections (untreated and treated with enoxaparin) labeling claudin-4 with ab15104 at 1/200. ab7090, anti-rabbit immunoglobulin G conjugated to horseradish peroxidase (HRP) at 1/300 was used as the secondary antibody. ab64238 at 1/50 was used to develop the histological signal.

    Antigen retrieval was performed by incubating the sections for 10 minutes at 97°C in 1 mM EDTA buffer, pH 8.0 or 10 mM citrate buffer, pH 6.0.

    Control, C; untreated colitis, DSS; oral enoxaparin, OE; intraperitoneal injection of enoxaparin, IPE.

  • Immuno-staining of formalin–fixed and paraffin-embedded liver sections with ab64238. Secontions were obtained from control (saline) and treated with CCl4+CT-N and CCl4+AD-N (left) and antibody control (right). 

  • Immunohistochemical analysis of mouse lung. Samples were fixed in 10% buffered formalin, paraffinized and sliced at 1.5 µm thick. Antigen retrieval was performed using ab64214 for the deparaffinized slices. Sections were blocked with 2% normal goat serum, PBS(-) and 0.1% Tween20. They were then incubated with the primary antibodies for 1 hour at 4°C and with secondary antibodies for 30 minutes at room temperature. The avidin-biotin-peroxidase complex method with peroxidase streptavidin and the DAB substrate kit ab64238 was performed.

    (A) A resected lung from a mouse sacrificed 28 days after SCL injection.

    (B) Haemotoxylin and Eosin staining showing the tumor composed of a central area with necrosis and a peripheral zone filled with SCLs.

    (C) Immunohistochemical staining for MMP-14 showing positive expression of MMP-14 in the peripheral zone of the tumor and negative in central zone.

    (D) Immunohistochemical staining weak staining of MMP-2 in the peripheral zone. 

References

This product has been referenced in:
  • Dan C  et al. HNF1B expression regulates ECI2 gene expression, potentially serving a role in prostate cancer progression. Oncol Lett 17:1094-1100 (2019). Read more (PubMed: 30655870) »
  • Vasconcelos ACU  et al. A symptomatic swelling of the upper lip. Oral Surg Oral Med Oral Pathol Oral Radiol 125:107-111 (2018). Read more (PubMed: 29248422) »
See all 43 Publications for this product

Customer reviews and Q&As

1-10 of 21 Abreviews or Q&A

Abreviews
This DAB substrate kit (ab94665) is easy to use, and the quality is good.


Liyong Zhang

Abcam user community

Verified customer

Submitted Aug 15 2017

Abreview for DAB substrate kit

Excellent Excellent 5/5 (Ease of Use)
Abreviews
This kit is streamlined to use at ease. The results is fantastic.


Liyong Zhang

Abcam user community

Verified customer

Submitted Aug 15 2017

IHC-P Mouse pancreas

Excellent Excellent 5/5 (Ease of Use)
Abreviews
Ki67 IHC staining on mouse pancreas, blocking with 10% BSA, 1 hr ab incubation RT

Abcam user community

Verified customer

Submitted Aug 16 2016

Answer

Thank you for contacting us.
The kit will work with the rabbit and mouse primary antibodies.
You can try citrate buffer pH for a start and Tris/EDTA in case the staining does not work. Please check the following for protocol and troubleshooting of;
https://www.abcam.com/index.html?pageconfig=resource&rid=11384
https://www.abcam.com/index.html?pageconfig=resource&rid=11467
I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.
Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

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Answer

Thank you for contacting us.
Heat mediated antigen retrieval method with microwave or pressure cooker is recommended for this ab.
The suitable detection system depends on the substrate you want to use. e.g. with DAB or AEC, we have following products;
https://www.abcam.com/DAB-Enhancer-ab675.html
https://www.abcam.com/DAB-Substrate-Kit-ab64238.html
https://www.abcam.com/AEC-Substrate-System-Ready-to-Use-ab64252.html
I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.
Free Rabbit monoclonal antibody with any purchase of a primary antibody, while stocks last! Quote “RABMAB-XBSMG” in your next primary antibody order. For more information, visit the following link: https://www.abcam.com/index.html?pageconfig=resource&rid=15447

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Answer

Thank you for contacting us again.



ab1002 Anti-Rabies Virus antibody looks very promising as it has been established for the use in IHC-P and should detect a wide range of rabies strains. As a starting dilution the customer can try a 5 ug/ml dilution incubated overnight at 4C. However, it may be that the actual optimum dilution may be lower or higher, but this depends on her samples. Another point is that I would strongly recommend a heat induced Antigen Retrieval if her slides are fixed with formalin.


https://www.abcam.com/index.html?datasheet=1002 (or use the following: https://www.abcam.com/index.html?datasheet=1002).


A matching secondary antibody would be Goat polyclonal Secondary Antibody to Mouse IgG2a - heavy chain (Biotin), pre-adsorbed (ab98696)


https://www.abcam.com/index.html?datasheet=98696 (or use the following: https://www.abcam.com/index.html?datasheet=98696).


To be fair, I am not the biggest fan of ready- to- use solution (andmost of my colleagues here a neither), for the following reason: most of all antibodies need optimization, may it be the blocking step, the retrieval time or method, the antibody incubation buffer etc. With ready-mix solutions it almost impossible to adjust the experiment settings. Ready mix solution make sense if the antibody is used in a high -throughput lab (like 20 slides at once with a fixed protocol), but not if you have to "learn" IHC-P and have the freedom to adjust the protocol fore the perfect results. In addition, it's cheaper ;)


Of course, once the protocol is optimized, using ready mixed solution are a very time efficient and make life easier, but I wouldn't use them as a beginner.


All the buffers your customer needs (see attachment) are listed in our very detailed and easy to follow step-by-step our protocol. However, none of my advice is written in stone, so if your customer prefers the ready mix solutions, we are more than happy to sell them to her.


Something we cannot provide are the reagents for the de-paraffinization, I am afraid, but a company like Sigma should be able to help with that.


Pease sent the customer our protocol and my advice. I am more than happy to assist her to get her experiments running. And another thing: she shouldn't be shy to ask any question she may have about the protocol!

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

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Answer

Thank you for contacting us.

Here is the list with the requested items. However, please be aware that this is not a technical inquiry as all item could be found by using the search application on our website under the "reagents" category. Please let us know if you need help on navigating our website so you can help your customers faster than relying on us.


Regarding the Rabies antibody, I am afraid we do not have what your customer requests: All we have are mouse monoclonal antibodies against this virus, but none produced in goat. Please let me know if this expectable for her, so I can suggest the right primary antibody with the matching biotinylated secondary antibody.


https://www.abcam.com/index.html?datasheet=64214 (or use the following: https://www.abcam.com/index.html?datasheet=64214).

https://www.abcam.com/index.html?datasheet=64216 (or use the following: https://www.abcam.com/index.html?datasheet=64216).

https://www.abcam.com/index.html?datasheet=64238 (or use the following: https://www.abcam.com/index.html?datasheet=64238).

https://www.abcam.com/index.html?datasheet=64269 (or use the following: https://www.abcam.com/index.html?datasheet=64269).

https://www.abcam.com/index.html?datasheet=94666 (or use the following: https://www.abcam.com/index.html?datasheet=94666).

https://www.abcam.com/index.html?datasheet=128988 (or use the following: https://www.abcam.com/index.html?datasheet=128988).

https://www.abcam.com/index.html?datasheet=128990 (or use the following: https://www.abcam.com/index.html?datasheet=128990).



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

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Answer

Thank you for contacting us. You are correct, these are the same product, just different sizes. The appropriate size will depend on how many slides you anticipate needing to stain. If you prepare the substrate/chromogen mixture as suggested in the protocol and apply it to individual slides with a pipettor, you will have enough solution in the 60ml kit to stain about 600 sections, assuming 100ul of mixture per slide, and over twice that many for the 125ml kit.

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Answer

Thank you for your reply.

I looked over the protocol that you sent and it looks to be OK with me. I had a colleague look at it aswell and they thought it was good too.

If there is anything else I can help you with, or if you have any further questions, then please let me know.

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Answer

Thank you for your reply and for providing those details.

I have now emailed you a proforma regarding the purchase of ab46667 and ab64238.

As for detailed step by step protocols, we do not have any that would meet your requirements, however the link below has many detailed protocols that would be useful to you:

http://ihcworld.com/protocol_database.htm#Histo-IHC



Please let me know if there is anything else I can help you with.

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1-10 of 21 Abreviews or Q&A

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