Biotinylation specifically refers to the process of attaching biotin to proteins or other macromolecules, including nucleic acid, functional groups and residues, in which antibodies are one of the most widely used targets.
Approaches of Antibody Biotinylation
Mainly two approaches are applied to create biotinylated antibodies, one of which is the avidin-biotin complex (ABC), and the other is the labeled streptavidin-biotin (LSAB).
l Avidin-biotin complex
The ABC method is broadly used for staining due to its modular and versatile with high sensitivity and low background. Most of the time, the ABC method is the first choice unless the avidin-biotin-enzyme complex is too large to pass through the tissue. There are some nonnegligible shortcomings also, such as nonspecific labeling caused by endogenous biotin and lower tissue penetration caused by large ABC complexes.
l Labeled streptavidin-biotin
The LSAB method, on the contrary, generates smaller complexes that allow better tissue penetration and higher sensitivity. Alternatives to avidin can be used in this method to reduce background noise and further sensitivity.
Based on the strongest known non-covalent interaction between biotin and avidin (or streptavidin), the process of biotinylation is completed fast without affecting the original function of the antibody. And the protein-ligand interaction of biotinylated antibodies is unaltered by extremes of pH, high salt concentrations, denaturant agents, temperature, and organic solvents.
in vivoin vitro
Biotinylated antibodies are designed to detect low-abundance proteins. Biotinylation also contributes to non-radiative labeling methods of proteins and protein purification, with increasing applications in medical and industrial fields, including biosensors, diagnosis, proximity assay, and drug screening.
A variety of biotinylated reagents allow different functional group specificities and different solubility characteristics, which substantially expands the range of applications for avidin-biotin chemistry research.