Avidin-biotin affinity

Numerous studies on bead bio chips were therefore based on magnetic [16,17], glass or silica [18-20], and polystyrene beads [21]. The DNA immobilization chemistry of those beads could be very different, from the classical avidin/biotin affinity reaction [17,22] (Fig. 2B) to the disulfide bridging onto thiol modified silica [19] (Fig. 5A), the thiocyanate reaction onto amino-terminated latex beads [21] (Fig. 5B), and finally the hybridization-based immobilization of poly(A)-tagged probes onto poly(T)-bearing magnetic beads [16] (Fig. 5C). 

The avidin-biotin complex, known for its extremely high affinity (Green, 1975), has been studied experimentally more extensively than most other protein-ligand systems. The adhesion forces between avidin and biotin have been measured directly by AFM experiments (Florin et al., 1994 Moy et al., 1994b Moy et al., 1994a). SMD simulations were performed on the entire tetramer of avidin with four biotins bound to investigate the microscopic detail of nnbinding of biotin from avidin (Izrailev et al., 1997). 

In another example, ligands can be biotinylated with a cleavable biotinylation reagent and then incubated with receptor molecules. The resulting complex can be isolated by affinity chromatography on immobilized (strept)avidin. Final purification of the ligand-receptor can be accomplished by cleaving the biotin modification sites while the complex is still bound to the support. The receptor complex thus can be eluted from the column without the usual harsh conditions required to break the avidin-biotin interaction. 

Figure 16.1 The general design of an ICAT reagent consists of a biotinylation compound with a spacer arm containing stable isotope substitutions. The reactive group is used to label proteins or peptides at particular functional groups and the biotin affinity tag is used to isolate labeled molecules using immobilized (strept)avidin.

More detailed discussions of avidin-biotin systems as well as the process of adding a biotin affinity group to proteins, nucleic acids, and other biomolecules can be found in Chapters 11 and 23. 

Morag, E., Bayer, E.A., and Wilchek, M. (1996) Immobilized nitro-avidin and nitro-streptavidin as reusable affinity matrices for application in avidin-biotin technology. Anal. Biochem. 243(2), 257-263. 

Avidin-biotin complex (ABC) is based on the high affinity that streptavidin (from Streptomyces avidinii) and avidin (from chicken egg) have for biotin. Biotin is a naturally occurring vitamin. One mole avidin will bind four moles biotin. ABC method affords a several-fold higher antigen detectability than those achieved in the standard indirect method. 

Fig. 12 Schematic illustration of immobilization strategy using affinity reactions via avidin-biotin coupling. Biotinylated mixed monolayers improve the availability of the surface-bound avidin or streptavidin to biotin coupled probe biomolecules...

Q Yang, X-Y Liu, M Hara, P Lundahl, J Miyake. Quantitative affinity chromatographic studies of mitochondrial cytochrome c binding to bacterial photosynthetic reaction center, reconstituted in liposome membranes and immobilized by detergent dialysis and avidin-biotin binding. Anal Chem 280 94-102, 2000. 

Y Tanaka, S Terabe. Studies of enantioselectivities of avidin, avidin-biotin complex and streptavidin by affinity capillary electrophoresis. Chromatographia 49 489-495, 1999. 

A rapid avidin/biotin ELISA has been developed for the determination of bovine somatotropin in blood and milk (150). The method uses affinity-purified polyclonal antisera raised in rabbits to immobilize bovine somatotropin from... 

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