Solid-Phase Biopanning / Phage Display
Biotinylate the target ligand, couple it to Dynabeads® Streptavidin and then expose to the phage or random DNA library. You can then separate the positive binders from negatives by magnetic handling.
Biopanning with Dynabeads® offers many advantages over plates, including :
- Correct orientation of the binding domain by directed biotinylation of the ligand.
- Minimal conformational changes of the immobilised target.
- Increased surface area allows for panning of saturated phage solutions.
- A higher percentage of positive binders and fewer rounds of panning.
- Magnetic separation allows for automation high throughput panning.
Phage display library screening is a valuable tool for protein-protein interaction mapping.
Dynabeads® Streptavidin have also been used to generate antigen-specific monoclonal antibodies in vitro. Ligands are immobilized on the beads, and high-affinity binders selected and matured by using a hypermutating B-cell line with only four rounds of sorting (5).
Dynabeads® are widely used in automated devices for panning for drug candidates, and a number of companies worldwide are benefiting from the unique properties of the Dynabeads® Streptavidin.
- Nord K. et al. (2001). Recombinant human factor VIII-specific affinity ligands selected from phage-displayed combinatorial libraries of protein A. Eur. J. Biochem. 268:4269-4277.
- Biroccio A. et al. (2002). Selection of RNA aptamers that are specific and high-affinity ligands of the hepatitis C virus RNA-dependent RNA polymerase. J. Virol. 76(8):3688-3696.
- Legendre D. et al. (1999). Engineering a regulatable enzyme for homogenous immunoassays. Nature Biotech. 17:67-72.
- Lev A. et al. (2002). Isolation and characterization of human recombinant antibodies endowed with the antigen-specific, major histocompatibility complex-restricted specificity of T-cells directed toward the widely expressed tumor T-cell epitopes of the telomerase catalytic subunit. Cancer Res. 62(11):3184-3194
- Cumbers SJ. et al. (2002). Generation and iterative affinity maturation of antibodies in vitro using hypermutationg B-cell lines. Nat. Biotech. 20(11):1129-1134.
- Demartis S. et al. (1999). A strategy for the isolation of catalytic activities from repertoires of enzymes displayed on phage. J. Mol. Biol. 286:617-633.
- Pini A. et al. (1998). Design and use of a phage display library. J. Biol. Chem. 273(34):21769-21776.
- Cox JC. et al. (2002). Automated selection of aptamers against protein targets translated in vitro: from gene to aptamer. Nucleic Acids Res. 30(20):e108.
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