Exploring minimal biotinylation conditions for biosensor analysis using capture chips.
Using Biacore's new regenerateable streptavidin capture (CAP) sensor chips, we investigated a number of biotinylation conditions for target ligands. We explored standard amine as well as the less commonly used carboxyl biotinylation methods. We illustrate the time scales required for efficient biotinylation as well as the hazards of overbiotinylation. We evaluated a range of desalting methods, including spin columns, dialysis membranes, and filters. Finally, we tested the effects of common buffer components, such as Tris and glycerol, on the biotinylation process. Together, our results serve as a general guide of the steps to consider when minimally biotinylating a target ligand.
Papalia
Giuseppe
G
Center for Biomolecular Interaction Analysis, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.
Myszka
David
D
D000818:Animals; D019921:Biotinylation / Q000379:methods*; D024402:Carbonic Anhydrase II / Q000378:metabolism; D002417:Cattle; D008024:Ligands; D015203:Reproducibility of Results; D019809:Streptavidin / Q000378:metabolism; D020349:Surface Plasmon Resonance / Q000295:instrumentation / Q000379:methods*
D008024:Ligands; D019809:Streptavidin; D024402:Carbonic Anhydrase II