Overview

  • Product name
    Mouse Interferon gamma ELISPOT Kit
    See all Interferon gamma kits
  • Detection method
    Colorimetric
  • Sample type
    Cell culture supernatant, Cell culture extracts
  • Assay type
    Sandwich (qualitative)
  • Assay duration
    Multiple steps standard assay
  • Species reactivity
    Reacts with: Mouse
  • Product overview

    The ELISPOT assay is designed to enumerate cytokine producing cells in a single cell suspension. This method has the advantage of requiring a minimum of in-vitro manipulations allowing cytokine production analysis as close as possible to in-vivo conditions in a highly specific way. This technique is designed to determine the frequency of cytokine producing cells under a given stimulation, and the follow-up of such frequency during a treatment and/or a pathological state. Elispot assay constitutes an ideal tool in the TH1 / TH2 response, vaccine development, viral infection monitoring and treatment, cancerology, infectious diseases, autoimmune diseases and transplantation.


    Abcam Elispot assay is based on sandwich immuno-enzyme technology. Cell secreted cytokines or soluble molecules are captured by coated antibodies avoiding diffusion in supernatant, protease degradation or binding on soluble membrane receptors. After cell removal, the captured cytokines are revealed by tracer antibodies and appropriate conjugates.


    Principle


    After cell stimulation, locally produced cytokines are captured by a specific monoclonal antibody. After cell lysis, trapped cytokine molecules are revealed by a secondary biotinylated detection antibody, which is in turn recognised by streptavidin conjugated to alkaline phosphatase. PVDF-bottomed-well plates are then incubated with BCIP/NBT substrate. Colored "purple" spots indicate cytokine production by individual cells.


     


    Recognizes natural murine Interferon gamma.

  • Tested applications
    Suitable for: ELISpotmore details
  • Platform
    Microplate

Properties

  • Storage instructions
    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests 5 x 96 tests
    Bovine Serum Albumin 1 x 2g 1 x 2g
    IFNγ Biotinylated detection antibody 1 x 100µl 1 vial
    Mouse IFNγ Pre-coated 96-well PDVF-bottomed plates 2 units 5 units
    Ready-to-use BCIP/NBT substrate buffer 1 x 11ml 2 x 25ml
    Streptavidin - Alkaline Phosphatase conjugated 1 x 10µl 1 x 50µl
  • Research areas
  • Function
    Produced by lymphocytes activated by specific antigens or mitogens. IFN-gamma, in addition to having antiviral activity, has important immunoregulatory functions. It is a potent activator of macrophages, it has antiproliferative effects on transformed cells and it can potentiate the antiviral and antitumor effects of the type I interferons.
  • Tissue specificity
    Released primarily from activated T lymphocytes.
  • Involvement in disease
    In Caucasians, genetic variation in IFNG is associated with the risk of aplastic anemia (AA) [MIM:609135]. AA is a rare disease in which the reduction of the circulating blood cells results from damage to the stem cell pool in bone marrow. In most patients, the stem cell lesion is caused by an autoimmune attack. T-lymphocytes, activated by an endogenous or exogenous, and most often unknown antigenic stimulus, secrete cytokines, including IFN-gamma, which would in turn be able to suppress hematopoiesis.
  • Sequence similarities
    Belongs to the type II (or gamma) interferon family.
  • Post-translational
    modifications
    Proteolytic processing produces C-terminal heterogeneity, with proteins ending alternatively at Gly-150, Met-157 or Gly-161.
  • Cellular localization
    Secreted.
  • Information by UniProt
  • Alternative names
    • IFG
    • IFI
    • IFN gamma
    • IFN, immune
    • IFN-gamma
    • IFNG
    • IFNG_HUMAN
    • Immune interferon
    • Interferon gamma
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab64029 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
ELISpot Use at an assay dependent dilution.

Protocols

References

ab64029 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-6 of 6 Abreviews or Q&A

Question
Answer

This kit ships with whole 96 wells plates, not 12, 8-well strips.

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Answer



Unfortunately, this kit is a ELISpot assay dedicated to detect IFNg secreting cells and not appropriate for serum.

This product will work for your customer's samples:

https://www.abcam.com/index.html?datasheet=46081 (or use the following: https://www.abcam.com/index.html?datasheet=46081).


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Answer

Thank you for contacting us.

The collaborator lab who devolved this kit have suggested using the following stimulation protocol :
Pre-stimulation of splenocytes by ConA (0.5µg/ml) + murine IL-2 (2ng/ml) 24h (or 48h), then addition of PMA (1ng/ml) and iono (500ng/ml).

I am sure with proper stimulation the kit ab64029 will be able to detect the high intensity spots.

We have received no complaint for ab46081 and the kit is optimized with the mentioned dilution of HRP. So I am sorry we are not aware if the recommended dilution is low. We will indeed investigate it if we get more complaints.

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 your email.

Peter is in feedback department. They do not deal with technical inquiries. I have been assigned your case and I will be following up now on. I have checked your previous contacts with my colleagues it seems you have problems with multiple kits. However I will be following this as a fresh complaint so please provide the requested information.

You have kindly submitted complaint for ab64029, which is ELISPOT kit however the protocol you have submitted as attachment is for ab46081. Could you clarify which kit to focus for this complaint?

The antibody in kit ab64029 is biotinylated with AP-streptavidin conjugated. It is not a HRP conjugated antibody so could you specify exactly what procedure you followed.

Am I right in saying the protocol of ab64029 and ab46081 were mixed without realizing different components. I might be wrong could you specify the details.

Many thanks for your cooperation in advance. I will look forward to hearing from you soon.

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Question
Answer

Thank you for calling us.

I am very sorry that we had so many problems sending you the emails. I am happy that it seems to work now.

I apologize also that our phone call got cut. Please do not hesitate to call back.

As we discussed on the phone, we would be pleased to look through the protocol to see whether we could suggest some optimisation tips. I amattaching thereforeour questionnaire so that we can gather further information regarding the samples tested and the protocol used. Please describe well the sample preparation. 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 look forward to receiving your reply.

Also, the catalogue numberof the mouse IL-4 ELISA Kit is ab100710. https://www.abcam.com/index.html?datasheet=100710 (or use the following: https://www.abcam.com/index.html?datasheet=100710).

Please let us know if you have any question.

I wish you a great weekend!

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Question
Answer

Thank youfor your phone call yesterday.

I am very sorry for the error on our datasheet in regards to the plate format. I have investigated this issue with my colleagues, and indeed, the kit ab64029 comes onlywith one 96 well plate. we have updated now the datasheet.I am very sorry for this error and the confusion generated. Unfortunately we do not have this Elispot kit with stripes.

I will be happy to send youan additional kit free of charge, so youcan perform more experiments.Unfortunately I do not know what I could do else to help you.

Maybe you can try to group the experiences? Also, it might be possible to use the free wells in the plate, even though the plate has been incubated. As said though on the telephone, I would not have recommended this. Indeed, this is not the intended use and it might not work. If you can include however the appropriate controls, you could assess whether it still works.

I am sorry for not being able to be more helpful. Please let me know if you have any questions or concerns and if you accept the free kit.

I am looking forward to hear back from you.

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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