With a dissociation constant (Kd) on the order of ≈10−14 mol/L,[1] the binding of biotin to streptavidin is one of the strongest non-covalent interactions known in nature.

Streptavidin is used extensively in molecular biology and bionanotechnology due to the streptavidin-biotin complex's resistance to organic solvents, denaturants (e.g. guanidinium chloride), detergents (e.g. SDS, Triton X-100), proteolytic enzymes, and extremes of temperature and pH.

The numerous crystal structures of the streptavidin-biotin complex have shed light on the origins of the remarkable affinity.

Most attempts at mutating streptavidin result in a lowered biotin-binding affinity, which is to be expected in such a highly optimized system.

However, a recently engineered mutant of streptavidin, named traptavidin, was found to have more than ten-fold slower biotin dissociation, in addition to higher thermal and mechanical stability.

Biotin-binding affinity can be impaired by chemical labeling of streptavidin, such as with amine-reactive fluorophores; flavidin is a streptavidin mutant without lysine side-chains, which retains good biotin binding characteristics after such fluorescent dye labeling where the dye couples to the amino terminus.

Harsh conditions are needed to break the streptavidin-biotin interaction, which often denatures the protein of interest being purified.

Streptavidin has also been used in the developing field of Nanobiotechnology, the use of biological molecules such as proteins or lipids to create nanoscale devices/structures.

[9] The tetrameric streptavidin has also been used as a hub around which other proteins may be arranged, either by an affinity tag such as Strep-tag or AviTag or by genetic fusion to SpyTag.

[15] Applications of monovalent streptavidin have included fluorescent tracking of cell surface receptors, decorating DNA origami, and acting as a pointer to identify specific regions for cryo-electron microscopy.

Monomeric streptavidin versions have an affinity for biotin of 10−7mol/L 10−8mol/L and so are not ideal for labeling applications but are useful for purification, where reversibility is desirable.

When the different tetramers are mixed together, a covalent linkage occurs to enable higher number of biotin binding sites.

Because streptavidin lacks any carbohydrate modification and has a near-neutral pI, it has the advantage of much lower nonspecific binding than avidin.