Biomolecule labeling continues to be a cornerstone feature of many in vitro and in vivo biological experiments. Click Chemistry has recently emerged as a convenient, versatile, and reliable method for labeling a wide variety of molecules for applications ranging from biomarker isolation to assay development.
LI-COR now offers a portfolio of Click Chemistry reagents for copper-catalyzed and copper-free methods. These products offer researchers flexibility to choose the correct reagent for a diverse array of applications. LI-COR Click Chemistry reagents include IRDye® 800CW, IRDye 680RD, and IRDye 650 near-infrared fluorescent dyes labeled with DBCO, azide, or alkyne groups.
Click Chemistry utilizes pairs of reagents that exclusively react with each other and are effectively inert to naturally-occurring functional groups such as amines. Unlike affinity interactions such as streptavidin-biotin, Click Chemistry forges covalent bonds between the reacting partners to deliver stable bioconjugates.
Click Chemistry reactions can be categorized into two separate groups, copper-catalyzed or copper-free. Copper-catalyzed Click Chemistry is used for initiating reactions between azides and alkynes. These reactions are also known as Copper-Catalyzed Azide-Alkyne Cycloaddition (CuAAC). Although they initiate and accelerate Click Reactions, copper catalysts are cytotoxic and inappropriate for use in living systems.
Chemiluminescent Western blot detection of ERK2 using WesternSure Reagents on the C-DiGit Blot Scanner. NIH/3T3 lysate (10 µg – 1.2 ng) was resolved on a 10% Bis-Tris gel and transferred to Odyssey nitrocellulose. The membrane was probed with anti-ERK2 (mouse mAb, Santa Cruz) followed by detection with WesternSure HRP Goat anti-Mouse IgG and WesternSure ULTRA Chemiluminescent Substrate. The Glow-Writer Pen was used to annotate the visible Odyssey Protein Molecular Weight Marker for chemiluminescent imaging on the C-DiGit Blot Scanner.
With WesternSure ULTRA Chemiluminescent Substrate, picogram amounts of antigen can be detected. This substrate is optimized for detection using an imaging system or film.
For more sensitive detection – in the femtogram range – try WesternSure PREMIUM Chemiluminescent Substrate. The PREMIUM substrate is optimized for highly sensitive detection and long signal duration, making it ideal for use with imaging systems.
Note: As of October 6, 2014, WesternSure ULTRA Chemiluminescent Substrate is no longer available. You may, however, want to try WesternSure ECL Substrate – Dura-like sensitivity at a Pico price. The Glow-Writer Pen is also no longer available, but we think you will find the WesternSure Pen a great alternative!
The dibenzocyclooctyne group (DBCO) allows copper-free Click Chemistry to be done with live cells, whole organisms, and non-living samples. DBCO groups will preferentially and spontaneously label molecules containing azide groups (—N3). Within physiological temperature and pH ranges, the DBCO group does not react with amines or hydroxyls, which are naturally present in many biomolecules. Reaction of the DBCO group with the azide group is significantly faster than with the sulfhydryl group (—SH, thiol).