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Antibody Labeling Kits

Lightning-Link®

For labeling primary antibodies, proteins or peptides

 

Antibody and protein labeling

What is Lightning-Link®?

Lightning-Link® is an innovative technology that enables direct labeling of antibodies, proteins, peptides or other biomolecules for use in R&D applications, drug discovery and the development of diagnostic kits. 

Despite its apparent simplicity, the Lightning-Link® process is sophisticated and generates conjugates with performance characteristics identical to, or better than, those prepared with laborious multistep conjugation procedures. 

What are the benefits Lightning-Link®?

Features Benefits
Quick and easy to use Save time, no special knowledge required
No separation steps 100% recovery - no antibody/protein loss
Can be used in a wide range of applications Flexible
Freeze dried Ships at ambient temperature, long shelf-life
Fully scalable (10ug to 1g or more) Easy transfer from R&D to manufacturing
Stringently QC tested Consistent high quality, excellent batch-to-batch reproducibility
Large number of labels available Experimental flexibility
Reliable: nearly 300 references Successfully used in many fields of research

How does Lightning-Link® work?

The researcher simply pipettes the antibody or other biomolecule into a vial of lyophilized mixture containing the label of interest and incubates for either 3 hours (Lightning-Link range) or only 15 min (Lightning-Link Rapid range)

ll process diagram

Where can I find more information about Lightning-Link®?
  1. Guide to antibody labeling 
  2. Lightning-Link Webinar
  3. FAQs
  4. Contact us
     

    Custom labeling services 

    Are you looking to increase assay sensitivity, decrease background or increase stability? Do you require bulk quantities to suit your manufacturing or research needs?

    At Innova we can help with all your bioconjugation requirements, so if you can't find what you’re looking for from our range of products, or need something a bit different or at a larger scale, our range of custom services may be of interest to you. These include:

    lightning link -testimonials-smaller fonts-v2

    See what others have said here!

    Lightning-Link® conjugates are used in a large number of scientific applications including:
    We also have a range of educational videos available, please follow the links below:

    Which label should I use?

    The choice of label for your antibody primarily depends upon the application:

    Immunoassay Label
    Western Blotting
    • Enzymes (usually HRP or Alkaline Phosphatase)
    ELISA
    • Enzymes
    • Biotin/Streptavidin
    Immunofluorescence
    • Fluorescent dyes
    Immunohistochemistry
    • Enzymes
    • Biotin/Streptavidin
    Flow Cytometry
    • Fluorescent proteins
    • Fluorescent dyes
    • Tandem dyes

    Please see the table here for absorption/emission wavelengths of Lightning-Link® fluorescent dyes, proteins and tandems.

    What is the difference between Lightning-Link® and Lightning-Link® Rapid?

    Both Lightning-Link® and Lightning-Link® Rapid kits can generate conjugates in a hands-on time of typically 20-30 seconds for the entire procedure. However the conjugation and quenching incubation times are shorter for Lightning-Link® Rapid kits allowing the conjugates to be generated faster. The antibody considerations are exactly the same, as is the high quality of the final conjugate.

    What can I label using Lightning-Link?

    Lightning-Link® technology works by targeting free amine groups on your target. It can be used to label antibodies, peptides, proteins and any other molecules with free amine groups.

    For advice on labeling molecules other than antibodies please contact our technical support team.

    How pure does my antibody need to be?

    Your antibody must be purified because other molecules with free amine groups will interfere with the reaction resulting in a poor quality conjugate.

    A suitable method of purification should be selected:

    • Affinity purification - this is the preferred method as it will always result in a purified sample containing only the antibody of interest.
    • Protein A or G purification of serum - this will result in a purified sample of all IgG’s contained in the serum. You will have a purified IgG fraction but only a small percentage of this fraction will be the IgG of interest. This IgG fraction will label with our kits, although adjustments in conjugate dilutions will be required.
    • Protein A or G purification of tissue culture supernatant or ascites fluid - this method will generate purified antibodies equivalent to affinity purification.

    Other purification methods, such as ion exchange chromatography and ammonium sulphate precipitation, are low quality purification methods and should be avoided. Filtration using a 0.2/0.22 µm filter is a method of sterilization not purification, and is therefore not appropriate.

    Is my antibody in a suitable buffer?

    For recommended buffer conditions please see the below table:

    Buffer components
    pH 6.5-8.5
    Amine free buffer (e.g. MES, MOPS, HEPES, PBS) ü
    Non-buffering salts (e.g. sodium chloride) ü
    Chelating agents (e.g. EDTA) ü
    Sugars ü
    Glycerol <50%
    Thiomersal/Thimerosal x
    Merthiolate x
    Sodium Azide1,3 <0.1%
    BSA1,2 <0.1% 
    Gelatin1,2 <0.1% 
    Tris <50mM
    Glycine
    Proclin
    Borate buffer ü 
    Nucleophilic components (Primary amines e.g. amino acids or ethanolamine and thiols e.g. mercaptoethanol or DTT) x

     1 Please note that individually the concentrations shown should not affect the reaction. However in combination with additional compounds that are not recommended above a certain concentration, the reaction may be affected.

    2 If intending to use this kit for immunohistochemistry, it is recommended that there be no gelatin or BSA present.

    3 It is important to note that sodium azide is a known inhibitor of HRP and should be avoided.

    What can I do if my antibody formulation does not fit the requirements?

    If your antibody requires purification or the buffer is not compatible with our kits, we have developed an AbSelect™ purification kit range that allows you to quickly and simply purify your antibody and is fully compatible with the Lightning-Link® kits.

    The appropriate kit to use depends on your particular sample (species, buffer, contaminants, volume, etc.). We have designed a handy flow chart on the AbSelect™ webpage to help you select a kit, but feel free to contact us if you require further guidance. Please consult the individual kit protocols to see the antibody amount/volume suitable for each kit.

    If your antibody is already purified but its concentration is significantly lower than 1mg/ml, you can concentrate it using our Antibody Concentration and Clean Up Kit. This kit can also be used to remove low molecular weight contaminants such as azide, tris or glycine by carrying out a buffer exchange into the buffer supplied in the kit, which is fully compatible with Lightning-Link®.

    If your antibody contains BSA, you can now use our BSA removal kit to purify your antibody in a few simple steps. Please note this kit will also enable you to concentrate your antibody.

    NB: All the AbSelect kits will ONLY work with antibodies. They will not purify other molecules. The only exception is the concentration and clean up kit (861-0010), which will work with other molecules greater than 10kD.

    Do I need a wash or desalt step?

    One of the advantages of the Lightning-Link® technology compared to traditional labeling methods is that conjugate purification is not required. The Lightning-Link® reactions are highly efficient, and the kits have been optimized so that, provided the protocols are followed, there should be only a very low amount of free label left at the end of the conjugation. This remaining free label would have its reactive 'Lightning-Link®' groups blocked by the Quencher provided in the kit, and is washed away during the wash step of your application.

    How stable will my new conjugate be?

    Upon dissolution of the Lightning-Link® mixture with a solution of the antibody, proprietary chemicals in the mixture become activated. This results in the directional covalent bonding of the antibody to the label in a gentle and controlled process at near-neutral pH. The stability of your conjugate is dependent on your antibody and label of choice.

    Most Lightning-Link® conjugates can be stored for up to 18 months, undiluted at 4°C. The exception to this is tandem conjugates which are only stable for up to 3 months.

     

     
    Lightning-Link ProductsClick the individual products for further information, pricing and data.

    Enzyme Labeling Kits

    Biotin / Streptavidin Labeling Kits

    Fluorescence Labeling Kits

    Lightning-Link®*
    Lightning-Link® Rapid*
    Maximal Absorbance
    (nm)
    Maximal Emission
    (nm)
    AMCA
     
    352
    452
    DyLight® 350
     
    354
    432
    Atto 390
     
    388
    468
    DyLight® 405
     
    402
    428
    PerCP 718-0030 484 678
    PerCP/Cy5.5 763-0030 484 692
    DyLight® 488   322-0030 496 524
    Fluorescein 498
    532
    R-Phycoerythrin
    498,544,566**
    580
    PE/Texas Red 767-0030 498,544,566** 618
    PE/Atto594 768-0030 498,544,566** 632
    PE/Cy5 760-0030 498,544,566** 672
    PE/Cy5.5 761-0030 498,544,566** 700
    PE/Cy7 762-0030 498,544,566** 782
    Atto 488   350-0030 504 530
    B-Phycoerythrin
    546
    580
    Cyanine Dye 3 (Cy3) 340-0030 552 576
    Rhodamine
    555
    588
    DyLight® 550
     
    556
    584
    Atto 565
     
    570
    598
    DyLight® 594
     
    594
    629
    Texas Red®
     
    596
    616
    DyLight® 633
     
    628
    660
    Atto 633 353-0030 634 660
    FluoProbes647H 362-0030 650 684
    Cyanine Dye 5 (Cy5)
     
    652
    678
    Allophycocyanin
     
    652
    666
    APC/Cy5.5 764-0030 652 700
    APC/Cy7
     
    652
    790
    DyLight® 650
    656
    686
    Cyanine Dye 5.5 (Cy5.5)
     
    680
    705
    DyLight® 680
     
    686
    716
    Atto 700
     
    704
    724
    DyLight® 755
     
    756
    794
    DyLight® 800
     
    776
    798
    * The antibody labeling kits come in two formats - Lightning-Link® and Lightning-Link® Rapid, with incubation times of 3 hours and 15 minutes respectively.
    **(R-)PE has three maxima, and all can be used. The optimal will depend on the application.

    For a downloadable version of the fluorescence table click here>>

    Fluorochrome guide download PDF

    by InnovaBiosciences 




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