Overview

  • Product name
    In-Cell ELISA (ICE) Support Pack
    See all In cell elisa kits
  • Sample type
    Adherent cells, Suspension cells
  • Assay type
    Cell-based (quantitative)
  • Assay duration
    Multiple steps standard assay
  • Product overview

    ab111542 is for use with suspension, apoptotic/detaching cells and adherent cell lines. The pack contains five 96-well microplates, buffers and protocol to perform ICE with Abcam's ICE-validated antibodies. For an ICE assay, it is necessary to purchase both primary antibody(ies) and labeled secondary antibody(ies). Antibodies are sold separately, allowing customizing the target(s) of interest, method of detection and multiplexing.


     

    In-Cell ELISA uses quantitative immunocytochemistry to measure protein levels or post-translational modifications within cells. The cells are fixed to the bottom of a coated 96-well plate (provided). Targets of interest are detected by primary antibodies, which are in turn quantified with labeled secondary antibody(ies). Abcam offers highly-specific, well-characterized primary antibodies, IRDye®- or HRP-labeled anti-mouse and anti-rabbit secondary antibodies, as well as IRDye®-labeled isotype specific anti-mouse antibodies. By combining antibodies of different species or isotype and appropriate IR-labeled secondary antibodies, two color multiplexing can be achieved in the 800/700 channels. IR imaging and quantification is performed using a LI-COR® Odyssey® or Aerius® system. HRP-labeled complexes are developed and quantified colorimerically using spectrophotometer.


    Two protocols are available when using this kit. Protocol (1) suspension cells and cells likely to detach under experimental conditions (for example, adherent cells undergoing apoptosis readily detach from a culture plate). Protocol (2) - A second protocol is available for normal adherent cells. These protocols can be used with any of Abcam's ICE-validated antibodies. Specific scientific information, background and working concentration for each antibody are detailed in each antibody's corresponding datasheet.

     

  • Tested applications
    Suitable for: In-Cell ELISAmore details
  • Platform
    Microplate

Properties

  • Storage instructions
    Store at +4°C. Please refer to protocols.
  • Components 5 x 96 tests
    100X Triton X-100 1 x 1.25ml
    10X Blocking Solution 1 x 40ml
    10X Phosphate Buffered Saline (PBS) 1 x 250ml
    400X Tween-20 1 x 4ml
    96-well Assay Plate 5 units
    Janus Green Stain 1 x 30ml
    Plate Seals 5 units

Applications

Our Abpromise guarantee covers the use of ab111542 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
In-Cell ELISA Use at an assay dependent dilution.

Images

  • Cells are grown to ~80% confluency in a 96- or 384-well plate, a drug/other treatment is applied to stimulate a cellular response. The cells are then fixed and permeabilized, effectively "freezing" them. Primary antibodies are then added which bind to their intended targets within the mitochondria or other subcellular compartment. After incubation, the unbound primary antibodies are washed away and secondary antibodies are added. These secondaries are conjugated to either IRDyes® or to an enzyme label (HRP or AP) for the colorimetric versions of the assays. Unbound secondaries are washed away, reaction buffer is added for the colorimetric assays, and the signal is read on a suitable instrument for the kit type.
    » In-cell ELISA diagram in PDF format

Protocols

References

This product has been referenced in:
  • Wang RE  et al. Rational design of a Kv1.3 channel-blocking antibody as a selective immunosuppressant. Proc Natl Acad Sci U S A 113:11501-11506 (2016). Read more (PubMed: 27663736) »
See 1 Publication for this product

Customer reviews and Q&As

1-6 of 6 Abreviews or Q&A

Answer


1. ab126420, the components of the blocking buffer are proprietary, and the solution is a standard blocking solution.

2. ab111542: The blocking solution in the kit is the same as the 10X blocking solution sold separately (ab126587). It is casein based and the % of casein is proprietary.

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Answer

Thank you for your inquiry.

I am happy to confirm that ICE can be used for nuclear proteins without any deviations from the standard protocol.

Please have a look at this link where more information can be found:

https://www.abcam.com/index.html?pageconfig=resource&rid=14181

We also offer a in ICE (In-Cell ELISA) Support Pack that includes all the reagents needed and a comprehensive protocol booklet for adherent and suspension cells.

ab111542

https://www.abcam.com/index.html?datasheet=111542 (or use the following: https://www.abcam.com/index.html?datasheet=111542).

I hope this information is helpful and wish you good luck.

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Answer

I am very pleased to hear you would like to accept our offer and testab616 and ab15580in ICE. These codes will give you: 1 free primary antibody per code before the expiration date. To redeem this offer, please submit an Abreview for ICE for each antibody and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. Please remember that submission of the Abreview is sufficient for the discount code to become active.For more information on how to submit an Abreview, please visit the site: https://www.abcam.com/Abreviews.

Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code.

Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

The terms and conditions applicable to this offer can be found here: https://www.abcam.com/collaborationdiscount.

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Question

Thank you so much. That is very helpful and informative. We are very interested in this assay, and if I can get buy-in from my seniors, I will be ordering this system soon and working on developing this assay. I was wondering if you should also emphasize in your promotional literature that this system has a couple more positive traits:

1. It has the potential for high-throughput screening of cell lines. If you are generating a cell line that is other-than-antibody, which we are doing more than one of, this system appears to me to allow the ready identification AND quantification of expressing clones. This need is what drove me to discover this technology.

2. It can be used on other fluorescent plate reading systems. It may be that you have an agreement with LICOR, but I know that Protein Simple (Alpha Innotech) and Molecular Devices (Isocyte) also have fluorescent plate-reading instruments. We have an Isocyte (we also have a LICOR—I haven’t used it) and it has the adaptability of filter swap-in-out such that at least two channels could be read simultaneously. It may be that the advantages of the LICOR readers are such that they are just superior, but if labs have other fluorescent readers, they may prefer not to purchase a separate instrument.

Again, thank you for your response and the information. Just as New England Biolabs is my favorite company for just about all products it sells—by far, you are my favorite antibody company—by far. And your technical expertise and support are responsible for at least part of that opinion.

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Answer

Thank you very much for your feedback! We are happy to help answer any technical questions you may have about In-Cell ELISA or any of our other assay or products. Please let me know if you need any additional information or assistance.

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Answer

Thank you for contacting us. Yes, In-Cell ELISA can be performed on suspension cells. Our ICE Support Pack ab111542 has been optimized for use on both adherent and suspension cells and provides the attached protocol for performing ICE on Suspension Cells. I hope this helps, please let me know if you have any additional questions.

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Answer

Thank you for your inquiry.



Unfortunately, we don’t offer the MetAbArray service any longersince we are 100% dedicated to individual product/assay development.


All of the antibodies that went into that offering (and many more now) are available as individual antibodies that could be used with our In cell ELISA protocol.

In addition we are offering multiple ICE duplexing kits with more to come in the next few months (Akt/phopho Akt, p53/phospho p53).

Here is a list of the ICE products we are offering. Please note that we've added a few more since this was updated, so if the targets aren't listed you can search our catalogue or I'd be happy to help you further.

https://www.abcam.com/index.html?pageconfig=resource&rid=14181

Ifyouareinterested in screening services using this technology, that approach is now being offered by MetProf (www.metprof.com).

The majority of the antibodies used in this platform are from Abcam, though, so if you have specific questions about these products, do not hesitate to contact us again.

Have a great day.

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Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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