Overview

  • Product name
    FITC Conjugation Kit
    See all FITC kits
  • Product overview

    FITC Conjugation Kit ab102884 uses a simple and quick process to conjugate an antibody to FITC . It can also be used to conjugate other proteins or peptides.


    To conjugate an antibody to FITC using this kit:
    - add modifier to antibody and incubate for 3 hrs
    - add quencher and incubate for 30 mins
    The conjugated antibody can be used immediately in WB, ELISA, IHC etc. No further purification is required and 100% of the antibody is recovered for use.


    Learn about buffer compatibility below; for incompatible buffers and low antibody concentrations, use our rapid antibody purification and concentration kits. Use the FAQ to learn more about the technology, or about conjugating other proteins and peptides to FITC.

  • Notes

    Amount and volume of antibody for conjugation to FITC 

     Kit size   Recommended 
    amount of antibody
    1 
    Maximum 
    amount of antibody
    Maximum antibody 
    volume2
    30 µg   3 x 10 µg  3 x 20 µg 3 x 10 µL
    300 µg   3 x 100 µg  3 x 200 µg 3 x 100 µL
    1 mg   1 mg 2 mg 1 mL

    1 Using the maximum amount of antibody may result in less labelling per antibody.

    2 Ideal antibody concentration is 1mg/ml. 0.5 - 1 mg/ml can be used if the maximum antibody volume is not exceeded. Antibodies > 5mg/ml or < 0.5 mg/ml should be diluted /concentrated.

     

    Buffer Requirements for Conjugation

    Buffer should be pH 6.5-8.5.

    Compatible buffer constituents 
    If a concentration is shown, then the constituent should be no more than the concentration shown. If several constituents are close to the limit of acceptable concentration, then this can inhibit conjugation.

    50mM / 0.6% Tris1 0.1%/1% BSA2 50% glycerol
    0.1% sodium azide PBS Potassium phosphate
    Sodium chloride HEPES Sucrose
    Sodium citrate EDTA Trehalose

    1 Tris buffered saline is almost always ≤ 50 mM / 0.6%
    2 1% BSA gives lower quality conjugates, BSA can also interfere with the use of the conjugated antibody in tissue staining.


    Incompatible buffer constituents

    Thiomerosal Proclin Glycine
    Arginine Glutathione DTT

    If a constituent of the buffer containing your antibody or protein is not listed above, please check the FAQ or contact us.

    Only purified antibodies are suitable for use, ie. where other proteins, peptides, or amino acids are not present: antibodies in ascites fluid, serum or hybridoma culture 

  • Tested applications
    Suitable for: Conjugationmore details

Properties

Applications

Our Abpromise guarantee covers the use of ab102884 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
Conjugation Use at an assay dependent dilution.

Images

  • This illustration demonstrates a general procedure and will slightly vary dependent on the conjugate used.

Protocols

References

This product has been referenced in:
  • Li Q  et al. Megalin mediates plasma membrane to mitochondria cross-talk and regulates mitochondrial metabolism. Cell Mol Life Sci 75:4021-4040 (2018). Read more (PubMed: 29916093) »
  • Zhdanov DD  et al. Murine regulatory T cells induce death of effector T, B, and NK lymphocytes through a contact-independent mechanism involving telomerase suppression and telomere-associated senescence. Cell Immunol 331:146-160 (2018). Read more (PubMed: 29935763) »
See all 13 Publications for this product

Customer reviews and Q&As

1-10 of 47 Abreviews or Q&A

FITC Conjugated full length EpCAM Protein

Excellent Excellent 5/5 (Ease of Use)
Abreviews
FITC was succesfully Conjugated with Human EpCAM Protein. It was quite useful.

Abcam user community

Verified customer

Submitted Nov 02 2015

Question
Answer

There is no difference in sensitivity between the rapid (eg. ab188285) and the standard (eg. ab102884) labeling kits. The only difference between the two is the incubation time required for the reaction.

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Better results than with secondary antibody.

Excellent Average 3/5 (Ease of Use)
Abreviews
The kit was easy enough for me to hand this off to an undergraduate, albeit with some explanation of the directions. These were a tad bit confusing at first, and not really clear on the volume of antibody to add (they give you a couple options since the Ab concentration of course matters, but didn't make it clear they were different options).
Otherwise the kit work fantastically with some store bought, pure, concentrated antibody! It actually gave better results than our previous method of using a conjugated secondary antibody when used in Flow Cytometry after doing an Intracellular Stain.

Hans Wilms

Verified customer

Submitted Jun 03 2014

Answer


This antibody will detect the whole cells, and because the antiserum has not been absorbed (only Protein A purified) it may react with related microorganisms.


As this antibody is suitable for conjugation purpose according to the datasheet, I would like to sduggest your customer is having a look at our FITC conjugation kit.


Read More

Answer

Thank you for your enquiry.

The EasyLink conjugation kits are designed so that they can be used to label proteins other than antibodies. In this case you should be able to label the Fc fused protein in the same way as labeling an antibody. However as the size of their protein is likely different to that of an average antibody, please find advice we give in these situations below:

As the protocols provided were optimised for labeling IgGs, we would recommend to adjust the amount of material added to the label vial to allow for molecular weight difference. This should be done without changing the volume added to the vial, as this could affect the conjugation efficiency. As a rough guideline, we would recommend changing the amount of material proportionally to the size difference with IgGs. An average IgG is about 160kD, therefore for a target that is ½ the size of an antibody (about 80kD), add ½ as much to the vial.

Finally, please note the usual EasyLink requirements will still apply: the target should be purified, in a suitable amine free buffer and not contain any additives such as Azide, BSA, Tris or Glycine.

I hope that this helps. If you have any additional questions please do not hesitate to contact me.

Read More

Answer

Thank you for contacting us.

The labeling kit works by targeting free amines groups on your antibody. We therefore cannot guarantee that the label will not bind to the FAB area, however this would be the case for any other amine orientated labeling technology. Although this might seem like an issue , it does not mean that the FAB will become unreactive, and we see successful conjugations in the great majority of cases.

The amine binding would only become an issue if there is an amine group within the antibody binding site itself.

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 your inquiry and your interest in our products.

I will have to double-check with the lab regarding glycosylation of the antibodies since I do not have this information readily at hand. I will let you know once I have received this information.

For the EasyLink kits, there is a quenching step which removes any excess label and reduces background. You can find more information about the labeling process here: https://www.abcam.com/index.html?pageconfig=resource&rid=13148&source=pagetrap&viapagetrap=easylink

Additional FAQs regarding these kits can be found here: https://www.abcam.com/index.html?pageconfig=resource&rid=13156

I will email you a quote for the 4 products. Currently we are running a promotion where if you buy any primary, you can get a RabMAb for free! Just mention the code RABMAB-XBSMG on the order and the cost of the rabbit monoclonal antibody will be removed from the order.

I hope this information helps. I will be in touch again with more information regarding the antibodies you requested.

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Answer

Thank you for your interest in the FITC conjugation kit. Unfortunately, it will be difficult to achieve a consistent ratio of FITC : IgG without extensive testing of the kit to determine the appropriate conditions. We have not done this testing, as the kit was developed for ease of producing a conjugate that simply gives a bright signal, rather than producing a conjugate of a known ratio. Further, the components of the reagents are considered proprietary, which makes knowing where to start difficult. To do what you want, I think it will make more sense to work with a standard FITC conjugation protocol, using known reagents and quantities, and testing various combination of FITC and antibody.

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Question
Answer

Thank you for contacting us.

The fluorescein in ourKit is not simply fluorescein and it is not FITC either. Our fluorescein has been specially treated with proprietary Easy-Link chemistry to make the unique one-step conjugation possible.

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!
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Answer

Thank you for your inquiries.

I can confirm that additives in the buffer of the antibody will interfere with the labeling reaction.

We offer a kit that will concentrate the antibodyand also remove the additives:

ab102776

https://www.abcam.com/index.html?datasheet=102776 (or use the following: https://www.abcam.com/index.html?datasheet=102776).

We usually recommend to label 100ug of antibody with the kits you purchased to achieve an optimal result. This means that one vial of ab18005 is enough for only one conjugation reaction.

It is possible to only use 50ug, but this is not optimal.

After the labeling of the antibody we recommend to store it at 4C to avoid freeze /thaw cycles that will harm the conjugate-antibody-complex.

Sodium azide can be added, but only to conjugate that are NOT HRP. HRP enzyme activity is inhibited by sodium azide.

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

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1-10 of 47 Abreviews or Q&A

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

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