Biotin surfaces

Application Note:

This product comes either in the form of a biotin surface for activation with streptavidin or in the form of pre-adsorbed streptavidin. The PEG environment ensures exceptionally low background. Control streptavidin amount at the surface is important.

Biotin-streptavidin interaction is one of the most widely used in bioconjugation chemistry, owing to the strong affinity and high specificity of the interaction. The binding strength between biotin and streptavidin is about four orders of magnitude stronger than that typical of antigen-antibody interaction and is the strongest among all noncovalent interactions. The presence of four binding sites on each streptavidin molecule makes it possible to link together biotin-tagged molecules or biotin-tagged molecules to a biotin functionalized surface. The exceptionally strong interaction ensures the integrity of the linker under a wide range of experimental conditions. When this interaction is used for the immobilization of a biotin tagged protein molecule to a solid surface, the interaction is through the tag, thus minimizing disturbance to protein structure.

To take advantage of the biotin-streptavidin interaction for protein immobilization, we have developed a surface of high density biotin groups on an otherwise zero background poly-ethyleneglycol (PEG) brush, Figure 1. As a demonstration of the specificity and exceptional low background of this surface, we show in Figure 2 the adsorption of Cy3-labeled streptavidin on the biotin/PEG suface. The surface remains resistant to the adsorption of fibrinogen.

With the adsorption of streptavidin on the biotin/PEG surface, the unoccupied binding sites on the tetravalent streptavidin molecule can be used for the immobilization of biotin-tagged proteins, as illustrated in Figure 3 for the adsorption of biotinlated BSA. The streptavidin surface can be freshly prepared on the biotin/PEG surface to ensure optimal interaction. The surface remains resistant to the adsorption of a “sticky” protein, fibrinogen.

The biotin/streptavdin surface is available on standard microscope slides, cover slips, and silicon wafers. We also provide customer coating service for specific samples and devices. Our customers have successfully applied the biotin/PEG surfaces for a range of applications, including protein sensors, protein microarrays, single molecule spectroscopy, biological atomic force microscopy and other biophysical experiments.

	Fig. 1. Left: biotin groups tethered to the high-density PEG coating. Right: the immobilization of a biotinlated protein molecule to the biotin/PEG surface via the intermediate streptavidin. biotin-streptavidin
	Fig. 2. Left: the specific adsorption of Cy3-labeledto the biotin-PEG surface. Note that the biotin surface remains resistant to the non-specific adsorption of fibrinogen (middle). Right: an array Cy3-streptavidin. protein immobilization on biotin surface
	Fig. 3. Left: adsorption of biotin-conjugated BSA on the streptavidin/biotin/PEG surface. The immobilized BSA is detected by immonostaining; Right: the streptavidin / biotin / PEG surface resists the nonspecific adsorption of a sticky protein, fibrinogen