biotinylates

Biotinylates refers to the process of covalently attaching biotin to a molecule, typically a protein, nucleic acid, or small molecule. The resulting biotinylated product can bind with very high affinity to streptavidin or avidin, enabling robust detection, purification, and immobilization in a wide range of biological and analytical workflows.

There are two main approaches to biotinylation. Enzymatic biotinylation uses biotin ligase (BirA) to attach a single biotin to a specific lysine within a short peptide tag (AviTag), providing site-specific labeling. Chemical biotinylation relies on reactive biotin derivatives such as NHS-biotin or sulfo-NHS-biotin, which form covalent amide bonds with primary amines on lysine residues or the N-terminus, often resulting in multi-site labeling and less predictable modification patterns.

Common reagents include NHS-biotin, sulfo-NHS-biotin (water-soluble), and PEG-linked variants to adjust sterics or solubility. Desthiobiotin and related analogs enable reversible binding to streptavidin, facilitating elution under milder conditions. For nucleic acids, biotin can be introduced during synthesis with biotinylated primers or biotinylated nucleotides, enabling subsequent affinity capture or detection.

Applications of biotinylates span purification, detection, and localization. Biotinylated proteins can be captured on streptavidin-coated beads for pull-down assays or affinity enrichment in proteomics. Biotin labeling supports Western blot detection, immunostaining, and imaging, as well as cell-surface labeling to study membrane proteins. In research and diagnostics, the strong biotin–streptavidin interaction provides a versatile platform for signal amplification and modular assay design.

Considerations include choosing site-specific methods when functional integrity is critical, balancing labeling efficiency with potential disruption of activity, and selecting reversible versus permanent biotin conjugates to fit the experimental workflow.