Overview

Properties

  • Form
    Liquid
  • Storage instructions
    Store at room temperature.
  • Storage buffer
    pH: 6.00
    Constituent: Citrate Buffer
    Note: Buffer x10 concentrated.
  • Concentration information loading...
  • Research areas
  • Relevance
    Citrate buffer is used on formalin-fixed, paraffin-embedded tissue sections mounted on glass slides for target retrieval prior to immunohistochemistry (IHC) procedures. Target retrieval prior to IHC procedures obtains positive results, or results in an increase in staining intensity with many primary antibodies.

Applications

Our Abpromise guarantee covers the use of ab64214 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 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. 

     

     

     

  • Immunohistochemical analysis of liver tissue section labeling PCNA with ab18197 at 1/250. Antigen retrieval was performed with antigen retrieval buffer, ab64236. A and B, PCNA staining in right and left grafts 1 day after reperfusion. E and F, PCNA staining in right and left grafts 5 day after reperfusion.

    This image was produced using ab64236 (a higher concentration of ab64214 ). Therefore, we expect ab64214 to work as well when diluted to the required concentration. 

     

References

This product has been referenced in:
  • He Y  et al. Tumor immunohistochemistry and preoperative magnetic resonance imaging features predict local recurrence of giant cell tumor of bone following intralesional curettage. Oncol Lett 17:1425-1434 (2019). Read more (PubMed: 30675196) »
  • Dzidziguri D  et al. Determination of The Properties of Rat Brain Thermostable Protein Complex which Inhibit Cell Proliferation. Cell J 19:552-558 (2018). Read more (PubMed: 29105389) »
See all 6 Publications for this product

Customer reviews and Q&As

1-5 of 5 Abreviews or Q&A

Answer

Thank you for your reply.

Your protocol seems comprehensive, although I would strongly recommend including the antigen retrieval step since this has worked well for other customers. We would recommend citrate buffer pH 6, which we do sell as ab64214.

https://www.abcam.com/10x-citrate-buffer-ph-6-0-ab64214.html

If you're looking for an antigen retrieval protocol, we have some online as well.

https://www.abcam.com/index.html?pageconfig=resource&rid=11488

If you do try the antigen retrieval step and still see no staining, I would be happy to replace or refund the product for you.

I hope this information helps. Please contact us with any other questions.

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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.

We do not currently offer double staining IHC kits. Your staining conditions are challenging as, of the two common HRP chromogens, only DAB is alcohol insoluable. While of the common AP chromagens, only BCIP/NBT is insoluable. This means that to do double labeling you will have to do a sequential stain first using one detection system and then with the other.

I have collected a list, with links, of the reagents which you would need for this type of staining.

1) After de-parrafinizing and re-hydrating the samples, perform an antigen retrieval step treating with Cirate buffer pH6.0,https://www.abcam.com/10x-Citrate-Buffer-pH-6-0-ab64214.html, in the microwave or pressure cooker. After performing the antigen retrieval rinse in cold tap water for 10 minutes.

2) Rinse slides in TBST,https://www.abcam.com/20x-TBS-T-with-Tween-20-ab64250.html.

3) Performing a blocking step using protein block,https://www.abcam.com/Protein-Block-ab64226.html.

4) Incubate sections with your first primary antibody, for clarity I will assume the CD44 antibody, raised in mouse.

5) Wash with TBST.

6) Block endogenous peroxidases using aHydrogen Peroxide Blocking Reagent,https://www.abcam.com/Hydrogen-Peroxide-Blocking-Reagent-ab94666.html.

7) Incubate your tissue with a biotinylated secondary antibody, in this instance anti-Mouse IgG,https://www.abcam.com/Biotinylated-Goat-anti-Mouse-IgG-H-L-Ready-to-Use-ab64255.html.

8) Wash with TBST.

9) Detect your first antibody usingStreptavidin Peroxidase (Ready to Use)https://www.abcam.com/Streptavidin-Peroxidase-Ready-to-Use-ab64269.html. Note that this does not containa serum or BSA solution as diluent. Serum may contain biotin therefore competing Streptavidin binding with biotinylated secondary antibody, therefore reducing binding activity.

10) Apply your chromogen, DABsubtrate,https://www.abcam.com/DAB-Substrate-Kit-ab64238.html.

11) Wash with TBST.

12)Performing a second blocking step using protein block,https://www.abcam.com/Protein-Block-ab64226.html.

13)Wash with TBST.

14)Incubate sections with the your second primary antibody, this time ALDH1, raised in rabbit.

15)Wash with TBST.

16)Incubate your tissue with a biotinylated secondary antibody, in this instance anti-Rabbit IgG,https://www.abcam.com/Biotinylated-Goat-Anti-Rabbit-IgG-H-L-Ready-to-use-ab64256.html

17)Wash with TBST.

18)Detect your first antibody usingStreptavidin Alkaline Phosphatase (Ready to Use)https://www.abcam.com/Streptavidin-Alkaline-Phosphatase-Ready-to-Use-ab64268.html.

19) Apply your second chromogenAlkaline Phosphatase chromogen (BCIP/NBT) - Ready to Use,https://www.abcam.com/Alkaline-Phosphatase-chromogen-BCIP-NBT-Ready-to-Use-ab7468.html.

18) Wash with TBST, counterstain if desired.


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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Question
Answer

Thank you for your recent telephone enquiry. I can confirm that the concentration of citrate in the buffer is 30 mM. I hope this will be helpful to you. Should you have any further questions, please do not hesitate to contact us. Good luck with your research!

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Answer

Thank you for your enquiry. Unfortunately, the concentration of 10 X citrate buffer is proprietary and can not be released. However, we can suggest that you make a ten times dilution of the buffer for the antigen retrieval. Simple take 1ml of this solution and dilute with 9ml of deionized water (DW). I hope this information helps. If there is anything else that I can help you with, please do not hesitate to contact me.

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