概述

  • 产品名称Avidin偶联试剂盒
  • 产品概述

    Abcam's Avidin Conjugation Kit provides a simple and quick process to conjugate your primary antibodies with Avidin.

    The conjugated antibody can be used straight away in WB, ELISA, IHC etc

    Learn more about buffer compatibility, protein/secondary antibody conjugation and labelling chemistry in our FAQs.


     

  • 说明

    Amount and volume of antibody

    The volume in which the antibody is added should ideally be 4-10µl. Antibody concentrations in the range 1-4mg/ml are therefore ideal. However, concentrations and volumes outside these suggested limits have also yielded good conjugates. For each new antibody, optimization of the ratio of antibody to Avidin is often worthwhile.

    Amount and volume of antibody

    1. Before you add antibody to the Avidin mix, add 1µl of Modifier reagent for each 10µl of antibody to be labeled. Mix gently.
    • Remove the screw cap from the vial of Avidin mix and pipette the antibody sample (with added Modifier) directly onto the lyophilized material. Resuspend gently by withdrawing and re-dispensing the liquid once or twice using a pipette.

     

    • Place the cap back on the vial and leave the vial standing for 3 hours in the dark at room temperature (20-25°C). Alternatively, and sometimes more conveniently, conjugations can be set up and left overnight, as the longer incubation time has no negative effect on the conjugate.

     

    • After incubating for 3 hours (or more), add 1µl of Quencher reagent for every 10µl of antibody used. The conjugate can be used after 30 minutes.

     

    Storage of conjugates

    For any new conjugate, initial storage at 4°C is recommended. A preservative may be desirable for long-term storage. Other storage conditions (e.g. frozen at -70°C or stored at -20°C with 50% glycerol) may also be satisfactory. The best conditions for any particular conjugate must be determined by experimentation.

    Sample Buffer

    Ideally, the antibody to be labeled should be in 10-50mM amine-free buffer pH range 6.5 to 8.5. However, many buffers outside these limits of concentration and pH can be accommodated. Modest concentrations of Tris buffer are also tolerated.

    Amine-free buffers, including MES, MOPS, HEPES and phosphate are compatible if they are in the concentration range 10-50mM and have pH values in the range 6.5-8.5, as the addition of Modifier provides the conditions necessary for efficient conjugation. Common non-buffering salts (e.g. sodium chloride), chelating agents (e.g. EDTA), and sugars may be present, as they have no effect on conjugation efficiency. Azide (0.02-0.1%) has little or no effect. If the amine-free buffer is relatively concentrated and outside the pH range 6.6-8.5 you may need to add more Modifier for each 10µl of antibody. Excess Modifier is provided so that you can check the pH of the buffer after the addition of the modifier. Ideally the pH should be around 7.3-7.6, though efficient conjugation occurs anywhere between pH 6.8 and 7.8. Avoid buffer components that are nucleophilic, as these may react with the kit chemicals. If your buffer contains primary amines (e.g. amino acids, ethanolamine) and/or thiols (e.g. mercaptoethanol, DTT), you should consider using our Concentration kits (Ab102776 or Ab102778).

    (Note: Unusually, for an amine, Tris has little effect on conjugation efficiency as long as the concentration is 20mM or less).

     

    Disclaimer

    Labeling of the antibody will not work if the conjugation blocks the active paratope. This situation is rare.

     

    The antibody to be labelled should be purified, in an appropriate buffer for conjugation and at a suitable concentration, as described in the protocol booklet. If not, consider using our antibody purification and concentration kits.

  • 经测试应用适用于: Conjugationmore details

性能

    应用

    Our Abpromise guarantee covers the use of ab102862 in the following tested applications.

    The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

    应用 Ab评论 说明
    Conjugation Use at an assay dependent dilution.

    Avidin Conjugation Kit 图像

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

    实验方案

    Avidin Conjugation Kit (ab102862)参考文献

    ab102862 has not yet been referenced specifically in any publications.

    Product Wall



    I can confirm that as long as there are free amines (and amines not conjugated to rhodamine) left in the bovine albumin, the conjugation of avidin to rhodamine coupled bovine albumin will be possible.

    The labeling chemistry targets p...

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    I can confirm that the quencher in the kit makes sure the conjugation has entirely finished before the conjugate is used further in an application. It will not interfere with the binding of the avidin-conjugated antibody to biotin.

    One of th...

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    Vielen Dank für Ihren Anruf.

    Wie versprochen haben ich das Labor, das den Kreatininantikörper ab30719 herstellt, kontaktiert, und wir können Ihnen bestätigen, dass der Antikörper sowohl freies (ungebundenes) als auch Carrierprotein-konjugier...

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    Thank you for your enquiry. The Easylink conjugation kits, including ab102862 EasyLink Avidin Conjugationkit, are primarily designed to label purified antibodies. However, because they work by targeting available amines they can be used to label other ...

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    Hello! I have more information regarding the buffers below: You can elute the antibody with citrate. Citrate is compatable with our LL kits. If you are using a protein A or G column you can use 10mM citrate buffer pH 3. If you are...

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    Thank you for contacting us. I have received additional advice from the lab regarding your question: It is possible to increase the level of antibody in the reaction to favor the addition of fewer avidins per antibody. If one were to add...

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    Thank you for your reply. The kit has been optimized for use with 10 ug of antibody. You could use more antibody (e.g. 20 ug), but ideally you would need to keep the volume as stated in the protocol at 10 ul. However, your labeling efficie...

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