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What about the amplification afforded by secondary indirect detection? |
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In many cases the ‘amplification’ of the indirect method may actually be illusory, as the primary antibody dissociates from the antigen during the incubation with the secondary reagent and during the following wash steps, thus what is actually being amplified is a diminishing amount of primary antibody. Therefore, in many cases the same level is obtained with direct detection, as shown by the following: |
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Western blotting data comparing directly vs indirectly labelled antibodies |
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Directly Labelled Antibodies |
Indirectly Labelled Antibodies |
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Primary antibody |
Goat anti-NQO1 |
Goat anti-NQO1 |
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Target |
Quinone reductase |
Quinone reductase |
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Sample lysate |
Human kidney |
Human kidney |
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Primary antibody working concentration |
0.00425 µg/mL |
0.1 µg/mL |
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Secondary antibody used |
None (direct conjugation) |
Yes |
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Exposure time (minutes) |
10 |
10 |
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Primary antibody source |
Everest Biotech, Cat. No.: EB05370 |
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Western blot analysis |
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Products |
Product Description
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Lightning-Link® Antibody and Protein Labelling Products |
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Lightning-Link the World’s easiest to use antibody and protein labelling kits. |
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Direct labelling (conjugation) of primary antibodies offers significant benefits over indirect detection using secondary antibodies. For instance, direct antibody labelling: |
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Circumvents problems of cross over and non-specific binding of secondary reagents. |
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Reduces the number of incubations and tedious wash steps. |
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Delivers a unique opportunity to study multi-protein complexes using antibodies generated from either the same or different species. |
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The Lightning-Link process - simply add purified antibody, protein or peptide to the lyophilised mixture |
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What Makes Lightning-Link Unique? |
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Easy to Use: |
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Hands on time of only 30 seconds |
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Conjugation reaction takes ca 3 hours at 20-25°C |
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No column separation steps are required to remove unbound labels |
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Molecules to be labelled only need to have a free amine group for efficient conjugation |
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Easy on your protein: |
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Conjugation is carried at physiological pH |
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Works with phosphate, HEPES, MES, MOPS and other amine free buffers |
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Reliable Results: |
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As the entire conjugation reaction is carried out in one tube, antibody recovery is 100% and labelling efficiency is very high |
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Reagents used in Lightning-Link are designed to deactivate and are completely benign after the labelling reaction and will not affect downstream experiments |
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Labelled antibody can be used directly in Western Blotting (WB), Enzyme Immunoassays (EIA's), Immunohistochemistry (IHC), Flow Cytometry Analysis and other applications which require labelled antibodies or other biomolecules |
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Convenient Pack Sizes: |
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All Lightning-Link Labelling kits are available in a number of convenient packs. The majority of kits are available in two or more of the following pack sizes: |
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10µg or 20µg per reaction, 3 reactions per kit
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100µg, 1 reaction per kit |
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100µg or 200µg per reaction, 3 reactions per kit
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1mg or 2mg per reaction, 1 reaction per kit
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Lightning-Link® Rapid Antibody and Protein Labelling Products |
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Lightning-Link Rapid the World’s fastest and easiest to use antibody and protein labelling kits. |
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The Lightning-Link Rapid Antibody Labelling kits offer all the same advantages as standard Lightning-Link kits, but now the labelled antibody is ready to use in just 20 minutes. |
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The Lightning-Link Rapid process - simply add purified antibody, protein or peptide to the lyophilised mixture and in 20 minutes, the antibody, protein or peptide is labelled and ready to use:
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Lightning-Link Rapid is currently available with the following labels and kit sizes: |
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Available Labels: |
Kit Sizes: |
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Atto390, 488, 565, 633, 700 |
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Cy3, Cy5, Cy5.5 |
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DyLight® 350, 405, 488, 550, 594, 633, 650, 680, 755 & 800 |
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FluoProbes 647H |
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AMCA, Fluorescein, Rhodamine, Texas Red®
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10µg or 20µg per reaction, 3 reactions per kit |
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100µg or 200µg per reaction, 3 reactions per kit |
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1mg or 2mg per reaction, 1 reaction per kit |
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imm-Link™ Immunogen Linking Kits |
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imm-Link Immunogen Linking Kits simplify the conjugation of carrier proteins to haptens: |
BSA, Ovalbumin and Keyhole Limpet Hemocyanin (KLH) stimulate a large immune response and are typically used as carrier proteins to generate antibodies against small antigens (peptides or organic compounds). For success the antigen, normally referred to as a hapten, needs to be covalently linked to the carrier protein.
Specific imm-Link Immunogen Linking Kits are designed for haptens with either sulfhydryl, carboxyl or amines functional groups which are conjugated to a carrier protein simply by adding a solution of the hapten to a proprietary lyophilised mixture containing the carrier protein and all the required conjugation chemistry. The hands-on time to set up the conjugation reaction is typically 20-30 seconds.
Following successful conjugation the conjugate is dialysed using the supplied dialysis cartridge to remove unwanted by products. The hapten-carrier protein conjugate can then be easily recovered and is ready for use. |
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All the imm-Link Immunogen Linking Kits are simply to use: |
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What Makes imm-Link Immunogen Linking Kits Unique? |
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Easy to Use: |
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All components supplied in one kit. |
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Wide range of chemistries. |
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Hands on time to set up conjugation reaction is 20-30 seconds. |
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Supplied dialysis cartridge ensures conjugates are recovered in high yield. |
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Available in 9 combinations of carrier proteins and conjugation chemistries: |
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Three Carrier Proteins: |
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BSA |
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Ovalbumin |
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Keyhole Limpet Hemocyanin (KLH) |
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Three Conjugation Chemistries: |
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imm-Link Sulfhydryl Immunogen Kit covalently binds the carrier protein to cysteines or free sulfhydryl groups (R-SH) on the hapten |
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imm-Link Carboxyl Immunogen Kit covalently binds the carrier protein to Glutamic Acid, Aspartic Acid or to free carboxyl groups (R-COOH) on the hapten |
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imm-Link Amine Immunogen Kit covalently binds the carrier protein to Lysines or free amines (R-NH2) on the hapten |
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Thunder-Link® PLUS Oligo Conjugation System
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Thunder-Link PLUS Oligo-Antibody Conjugation |
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Simplifies the generation of oligo-antibody conjugates |
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Antibody – oligonucleotide conjugates are becoming increasingly popular as a tool for biomarker detection. Typically oligonucleotide conjugates are used in immuno-PCR which combines the specificity of antibodies with the amplification power of immuno-PCR allowing a 10 to 1000 fold increase in sensitivity. Oligonucleotide conjugates also have the potential to be the platform tool in multiplexed protein diagnostic assays.
Thunder-Link PLUS enables simple and rapid conjugation of antibodies to oligonucleotides, with high recovery of materials and a superior clean-up procedure. The kit is quick and simple to use, overcoming time consuming and lengthy protocols associated with standard conjugation methods.
Thunder-Link® PLUS has been developed as a superior alternative to our original Thunder-Link oligo conjugation system, enabling even faster and more efficient conjugation.
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Features of Thunder-Link Oligonucleotide-Antibody Conjugation: |
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Only 30 minutes antibody and oligo activation |
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Fast oligo conjugation – now only 1 hour! |
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The single stranded oligo must be between 20 and 120 bases in length and contain a terminal amine group. Oligos may be either 3 or 5’ aminated.
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High antibody and oligo recovery |
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Robust and flexible clean-up procedure (now works with antibody fragments and other proteins) |
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Activated Labels |
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Maleimide Activated Labels |
Maleimide activated labels are designed to facilitate the conjugation of the desired label to either antibodies, proteins, peptides or ligands that contain a sulfhydryl (R-SH) group.
The reactive groups that have been added to the label allows conjugation reactions to be performed very efficiently at physiological pH for superior performance. |
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Convenient and Easy to Use: |
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Eight maleimide activated labels and one periodate activated label are available |
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Supplied as freeze dried formulations, therefore shipping on dry ice is unnecessary |
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Can be used to label proteins, peptide and ligands |
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Labelling performed at physiological pH |
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Available Maleimide Activated Labels: |
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Alkaline Phosphatase (AP)
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Allophycocyanin-XL |
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BSA |
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Horseradish Peroxidase (HRP) |
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Keyhole Limpet Hemocyanin (KLH) |
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Ovalbumin |
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R-Phycoerythrin (R-PE) |
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Streptavidin |
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Antibody Purification |
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Antibody Purification Systems
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The AbSelect range of antibody purification systems are cost effective and easy to use antibody purification systems which have been specifically designed to be compatible with a wide range of matrices. All the components of all the AbSelect™ kits are compatible with the Lightning-Link® Antibody Labelling kits, hence the purified antibodies can be immediately conjugated with the desired label. |
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AbSelect™ Antibody Purification System |
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Used to purify commerciallyavailable antibodies which often contain substances (e.g. BSA, glycine, tris, azide) that interfere in labelling reactions. The AbSelect™ Purification Kit quickly removes these contaminants. It can also be used to purify antibodies from crude samples such as ascites fluid or immune serum. |
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Convenient and Easy to Use: |
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Suitable for volumes of 100µL to 0.5mL |
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Up to 500µg of antibody can be purified in each run |
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AbSelect™ G Antibody Purification System |
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Used to purify commercially available antibodies which often contain substances (e.g. BSA, glycine, tris and azide) that interfere in labelling reactions with enzymes, biotin, streptavidin or fluorophores. The AbSelect™ G Purification System quickly removes these contaminants. It can also be used to purify antibodies from crude samples such as ascites fluid or immune serum. |
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Convenient and Easy to Use: |
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Suitable for volumes of 100µL to 0.5mL |
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Up to 500µg of antibody can be purified in each run |
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AbSelect™ G Serum Antibody Purification System |
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Antibodies generated from ascites fluid and serum are often supplied as crude formulations. Both ascites fluid and serum contain many substances that interfere with antibody labelling reactions. Successful labelling therefore requires antibodies to be purified prior to labelling. The AbSelect™ G Serum system is a fast and simple method to purify antibodies from these types of media. Protein G has a high affinity for the Fc regions of IgG molecules from a variety of species. |
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Convenient and Easy to Use: |
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Used to purify IgG from both ascites and serum |
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Upto 10mg of antibody can be purified in each run |
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AbSelect™ Mouse Antibody Purification System |
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Use to purify commercially available antibodies often contain substances (e.g. BSA, glycine, tris and azide) that interfere in labeling reactions with enzymes, biotin, streptavidin or fluorophores. The AbSelect™ Mouse Antibody Purification System quickly removes these contaminants. It can also be used to purify mouse antibodies from crude samples such as ascites fluid. |
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Convenient and Easy to Use: |
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Suitable for volumes of 100µL to 0.5mL |
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20µ to 150µg of antibody can be purified in each run |
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AbSelect™ Serum Antibody Purification System |
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Used to purify antibodies generated from ascites fluid and serum which are often supplied as crude formulations which contain many substances that interfere with antibody labelling reactions. |
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Convenient and Easy to Use: |
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Suitable for volumes of 4mL to 40mL of ascites fluid |
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Suitable volume for serum is species dependent |
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Upto 20mg of antibody can be purified in each run |
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AbSelect™ Mouse TCS Purification System |
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Used to purify mouse IgG fractions from hybridoma supernatants. The method involves capture of the antibody on the AbSelect™ Mouse TCS resin which has a high affinity for mouse IgG and the removal of unwanted substances using a simple wash procedure. The purified product is then eluted and neutralized. The AbSelect™ Mouse TCS Purification System is not suitable for use with antibodies from other species. It should be noted that the binding strength of Bovine IgG to the AbSelect™ Mouse resin is negligible. |
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Convenient and Easy to Use: |
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Suitable for volumes of 10mL to 25mL of tissue culture supernatant. |
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Upto 1.5mg of antibody can be purified in each run |
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AbSelect™ Tissue Culture Supernatant (TCS) Antibody Purification System |
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Usd to purify antibodies which are generated from hybridoma cell lines and supplied in tissue culture supernatant which contains many substances that interfere with antibody labelling reactions. |
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Convenient and Easy to Use: |
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Suitable for volumes of 10mL to 50mL of tissue culture supernatant. |
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Up to 5mg of antibody can be purified in each run |
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DyLight® is a registered trademark of Thermo Fisher Scientific Inc. and its subsidiaries. |
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Texas Red® is registered trademark of Life Technologies Corporation. |
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Products and Applications 
