Biotinylation is one of the most widely used bioconjugation techniques that covalently attaches a biotin tag to a protein, nucleic acid or other molecule for high-affinity binding with streptavidin reagents. Biotinylation is a rapid, specific reaction, and due to the small size of biotin, the process does not interfere with the natural function of the molecule. Biotin binds to streptavidin and avidin with an extremely high affinity, fast on-rate, and high specificity, and this binding reaction is exploited in many areas of biotechnology to isolate biotinylated (biotin-labeled) molecules of interest. Biotin binding with streptavidin and avidin resists extreme heat, pH, and proteolysis, making it possible to capture biotin molecules in a variety of environments. In addition, multiple biotin molecules can be conjugated to a target protein, which allows multiple streptavidin, avidin, or neutravidin protein molecules to bind and increases the detection sensitivity of the target protein. Biotinylated proteins are usually detected or purified with avidin conjugates in many protein research applications such as enzyme-linked immunosorbent assay (ELISA), western blot (WB), immunohistochemistry (IHC), immunoprecipitation (IP), and flow cytometry/fluorescence activated cell sorting (FACS).
A large number of biotinylation reagents are available in a wide range of possible labeling methods. The chemical reagents used for protein biotinylation consist of a biotin moiety, a spacer, and a reactive moiety. The biotin moiety has the valerate side chain, which plays an important role in the interaction with avidin. The terminal carboxyl group of the chain can be easily derivatized and conjugated to various reactive moieties and chemical structures, which allows various biotinylation reagents to be developed. The spacer has sufficient length to allow protein capture by the immobilized streptavidin. It is an important variable, which largely determine the properties of the biotinylated reagent and the derivatized protein, such as availability, solubility, and biotinylation reversibility. The spacer can also be cleaved by chemical or physical reagents to enable protein to release after capture. The reactive moiety is for the covalent binding of biotin to the target protein. The most common reactive groups include alkynes, azides, maleimides, hydrazine, N-hydroxysuccinimide (NHS) esters, etc.
Amerigo Scientific offers a wide range of biotinylation reagents for targeting specific functional groups or residues, such as primary amines, azides, sulfhydryls, carboxyls and carbohydrates. We also provide biotinylation reagents that combine biotin with single-molecular weight, discrete PEG linkers, and the reactive groups of these molecules allow many different types of conjugation reactions. Moreover, these water-soluble biotinylation products do not cause aggregation and precipitation of conjugates that often occur when hydrophobic biotinylated compounds are used.
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