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Agarose Avidin D (A-2010) is prepared by conjugating Avidin D to heat stable, cross-linked 4% agarose gel beads. To ensure minimal steric interference and low nonspecific binding, Avidin D is conjugated through a hydrophilic spacer arm. The procedure we have developed for coupling Avidin D to agarose optimally preserves the biotin binding activity of the Avidin D. Unlike cyanogen bromide coupling, our procedure does not produce conjugates which can be leached from the gel with solutes such as Tris buffer. Our procedure also does not generate charged groups on the gel that can bind proteins nonspecifically. Protein concentration is 2 mg Avidin D per ml of settled agarose beads. The product is supplied as a 1:1 suspension in buffer. The binding capacity is greater than 2 mg of biotinylated IgG per ml of settled beads. |