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Easy Antibody Conjugation

Info

Lightning-Link™ the world’s easiest to use antibody labeling kits. Simply add purified antibody to the lyophilised mixture.


Overview

Direct labeling of primary antibodies, offers significant benefits over secondary indirect detection. For instance, direct antibody labeling:

 

If you are new to the Lightning-Link™ kits, we invite you to:

 

Features

 

Benefits

 

Lightning-Link Antibody Conjugation Products

Lightning-Link™ is a one step antibody labeling method that requires only 30 seconds of hands on time, and as there are no separation steps antibody recovery is 100%. Find out more about these products:

 

Conjugation to Enzymes

Lightning-Link HRP (horseradish peroxidase)
Avoid high background as a result of secondary antibodies in your immunohistochemistry experiments – go direct!

Lightning-Link AP (alkaline phosphatase)
Alkaline phosphatase (AP) is a hydrolyase enzyme that is frequently conjugated to antibodies for use in immunoassays. PNPP is a popular colorimetric substrate for AP; the product of the reaction, p-nitrophenol, can be read at 405 nm. Fluorogenic substrates (e.g. 4-methylumbelliferyl phosphate; MUP) may also be employed.

Lightning-Link GOx (glucose oxidase)
Glucose oxidase is an enzyme produced and purified from Aspergillus niger which catalyses the oxidation of glucose with the release of hydrogen peroxide. Its molecular weight is 80 kDa, but it exists as a dimer of 160 kDa Measurement of glucose by GOx is used in the food industry, in fermentation and, most importantly, as the basis of biosensors for the diagnosis of diabetes.

 

Conjugation to Fluorescent Proteins

Lightning-Link APC (allophycocyanin)
Allophycocyanin is a large 105 kDa fluorescent phycobiliprotein commonly used in FACS analysis. It absorbs at 650 nm and emits at 662 nm.

Lightning-Link B-PE (B-phycoerythrin)
Useful for flow cytometry. B-Phycoerythrin absorbs strongly at about 545 nm (slightly yellowish green) and emits strongly at 572 nm (yellow). B-Phycoerythrin may also be less "sticky" than R-Phycoerythrin and contributes less to background signal due to non-specific binding in certain applications

Lightning-Link R-PE (R-phycoerythrin)
Useful for flow cytometry. R-Phycoerythrin (R-PE) is a fluorescent protein from the phycobiliprotein family, and is isolated from red algae. The absorbance spectrum of R-PE covers a broad range of excitation wavelengths, which provides an advantage for multi-color immunofluorescent staining or cell sorting. R-PE is one of the most intensely fluorescent phycobiliprotein having orange fluorescence. It is significantly brighter and more photostable than conventional organic fluorophores and it has a high quantum yield.

Lightning-Link PerCP (peridinin-chlorophyll-protein)
PerCP is a small 35 kDa phycobiliprotein isolated from red algae. It has a strong absorption peak at 482 nm and a secondary absorption peak at 442 nm. PerCP has a large stokes shift (195 nm) with its maximum emission peak at 677 nm. PerCP is commonly used for fluorescent immunolabeling, particularly in applications involving fluorescent-activated cell sorting.

 

Conjugation to Biotin, Streptavidin or Avidin

Lightning-Link Biotin Type A (for assays with labelled streptavidin, e. g. Strep-HRP)
The attachment of biotin to biomolecules is an important laboratory technique. Biotin binds to the tetrameric avidin proteins, including streptavidin and neutravidin, with exceptionally high affinity, and this interaction is exploited in various applications such as western blotting, immunohistochemistry and ELISA. The biotin in the kit has an extended linker to facilitate molecular interactions.

Lightning-Link Biotin has been optimised for two separate applications. Type A is intended for assays in which a streptavidin-labeled detection reagent will be used.

Lightning-Link Biotin Type B (for assays with immobilized streptavidin)
Type B is optimised for assays in which the biotinylated protein is captured by streptavidin immobilized on a surface (i. e. plates, nitrocellulose, magnetic beads etc).

Lightning-Link Avidin
Avidin is a 66 kDa tetrameric glycoprotein composed of four identical subunits, which has widespread applications due to its very high affinity to the vitamin biotin.

Lightning-Link Streptavidin
Streptavidin is a 53 kDa protein purified from Streptomyces avidinii, which has widespread applications due to its very high affinity to the vitamin biotin. Streptavidin is a tetrameric molecule, composed of 4 13 kDa monomers, each of which can bind a molecule of biotin.

 

Conjugation to Tandem Dyes

Lightning-Link Tandem APC/Cy5.5 (λabs: 633 nm, λfl: 694 nm)
APC/Cy5.5 is a tandem conjugate. The APC is excited at 633 nm and functions as an energy donor for the Cy5.5. Energy is transferred from the APC to the Cy5.5 via FRET (Fluorescence Resonance Energy Transfer). The Cy5.5 emits then the energy received from the APC in the form of long wavelength light at 694 nm.

Lightning-Link Tandem APC/Cy7 (λabs: 633 nm, λfl: 767 nm)
APC/Cy7 is a tandem conjugate. The APC is excited at 633 nm and functions as an energy donor for the Cy7. Energy is transferred from the APC to the Cy7 via FRET (Fluorescence Resonance Energy Transfer). The Cy7 emits the energy received from the APC in the form of long wavelength light at 767 nm.

Lightning-Link Tandem PE/Texas Red (λabs: 488 nm, λfl: 615 nm)
PE/Texas red is a tandem conjugate. The PE is excited at 488 nm and functions as an energy donor for the Texas red. Energy is transferred from the PE to the Texas red via energy resonance transfer. The Texas red emits the energy received from the PE in the form of long wavelength light at 615 nm.

Lightning-Link Tandem PE/Atto594 (λabs: 488 nm, λfl: 627 nm)
PE/Atto594 is a completely new tandem conjugate. The PE is excited at 488 nm and functions as an energy donor for the Atto594 fluorescent dye. Energy is transferred from the PE to the Atto594 dye via energy resonance transfer. The Atto594 emits the energy received from the PE in the form of long wavelength light at 627 nm.

Lightning-Link Tandem PE/Cy5 (λabs: 488 nm, λfl: 679 nm)
PE/Cy5 is a tandem conjugate. The PE is excited at 488 nm and functions as an energy donor for the Cy5. Energy is transferred from the PE to the Cy5 via energy resonance transfer. The Cy5 emits the energy received from the PE in the form of long wavelength light at 679 nm.

Lightning-Link Tandem PE/Cy5.5 (λabs: 488 nm, λfl: 695 nm)
PE/Cy5.5 is a tandem conjugate. The PE is excited at 490 nm and functions as an energy donor for the Cy5.5. Energy is transferred from the PE to the Cy5.5 via energy resonance transfer. The Cy5.5 emits the energy received from the PE in the form of long wavelength light at 695 nm.

Lightning-Link Tandem PE/Cy7 (λabs: 488 nm, λfl: 776 nm)
PE/Cy7 is a tandem conjugate. The PE is excited at 488 nm and functions as an energy donor for the Cy7. Energy is transferred from the PE to the Cy7 via energy resonance transfer. The Cy7 emits the energy received from the PE in the form of long wavelength light at 776 nm.

Lightning-Link Tandem PerCP/Cy5.5 (λabs: 482 nm, λfl: 700 nm)
PerCP/Cy5.5 is a tandem conjugate. The PerCP is excited at 482 nm and functions as an energy donor for the Cy5.5. Energy is transferred from the PerCP to the Cy5.5 via energy resonance transfer. The Cy5.5 emits the energy received from the PerCP in the form of long wavelength light at 700 nm.

 

Conjugation to Fluorescent Dyes

Lightning-Link AMCA (λabs: 346 nm, λfl: 442 nm)
Unlike standard AMCA labeling procedures, where a large molar excess of AMCA is employed, the Lightning-Link AMCA kit combines a low molar ratio of AMCA:antibody and high labeling efficiency to eliminate the need for purification steps. AMCA (aminomethylcoumarin acetate) is a blue fluorescent dye commonly used in fluorescence microscopy, arrays and in-situ hybridisation. It has an excitation wavelength of 345-350 nm and an emission wavelength of 440-460 nm.

Lightning-Link Fluorescein (λabs: 485 nm, λfl: 535 nm)
Fluorescein is a fluorophore widely used in biological analysis, with a pH dependent absorption and emission spectrum. In water it shows an absorption maximum at 494 nm and an emission maximum of 521 nm (in water).

Lightning-Link Rhodamine (λabs: 544 nm, λfl: 576 nm)
Rhodamine dyes are used extensively in biotechnology applications such as fluorescence microscopy, flow cytometry, fluorescence correlation spectroscopy and ELISA. Rhodamine has an emmission wavelength of 576 nm and an excitation wavelength of 544 nm.

Lightning-Link Cy3 (λabs: 550 nm, λfl: 570 nm)
Cyanine Dye 3 (Cy3) has a strong absorption peak at 550 nm and a secondary absorption peak at 520 nm. The maximum emission peak is at 570 nm, with an excitation coefficient of 1.5 x10^5/(cm M). Cy3 can be used as an alternative to Alexa Fluor 555, DY-550, Rhodamine Red-X and TAMRA [5(6)-Carboxytetramethylrhodamine].

Lightning-Link Texas Red (λabs: 595 nm, λfl: 615 nm)
Texas Red is a red fluorescent dye, used in histology, in fluorescence microscopy applications and in immunohistochemistry. It has a strong absorption at 595 nm, with a high fluorescence at 615 nm (extinction coefficient 8.0 x10^4 /(cm M) and high quantum yield.

Lightning-Link Cy5.5 (λabs: 643 nm, λfl: 667 nm)
Cyanine Dye 5.5 (Cy5.5) has a strong absorption peak at 675 nm. The maximum emission peak is at 694 nm, with an extinction coefficient of 2.5 x10^5/(cm M). Cy5.5 can be used as an alternative to Alexa Fluor 680, WellRed DP-3A, DY- 676 and LightCycler Red.

Lightning-Link Atto700 (λabs: 700 nm, λfl: 725 nm)
Atto700 is one of a new generation of fluorescent labels that emits in the red spectral region. It has a strong absorption peak at 690 nm, with an extinction coefficient of 1.2 x10^5/(cm M), and a maximum emission peak at 725 nm.

 

Tips for Antibody Purification

Download your Antibody Purification Guide (pdf, 574 kB).

 

Antibody Purification for Labeling Scheme

 

View all Innova Biosciences Conjugation Kits


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Antibody Purification KitsLightning-Link™ Frequently Asked Questions


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