Soybean agglutinin (SBA), Agarose bound

Composed of four subunits of approximately equal size, soybean agglutinin is a family of closely related isolectins. This glycoprotein has a molecular weight of about 120 kDa and an isoelectric point near pH 6.0. SBA preferentially binds to oligosaccharide structures with terminal α- or β-linked N-acetylgalactosamine, and to a lesser extent, galactose residues. Binding can be blocked by substitutions on penultimate sugars, such as fucose attached to the penultimate galactose in blood group B substance. An important application for SBA is the separation of pluripotent stem cells from human bone marrow. Cells fractionated by SBA do not produce graft vs host disease and can be used in bone marrow transplantation across histocompatibility barriers. Agarose bound* Soybean agglutinin is prepared using our affinity-purified lectins. Heat stable, cross-linked 4% agarose beads with a molecular weight exclusion limit of about 2x107 daltons are used as the solid-phase matrix to which the lectins are covalently coupled. The attachment of the lectins to the beads is carefully controlled to preserve lectin activity and minimize conformational changes of the bound lectins that might result in nonspecific ionic or hydrophobic interactions. The technique we have developed to couple lectins to agarose beads inserts a hydrophilic spacer arm between the lectin and the matrix. This coupling method provides several advantages over the traditional cyanogen bromide procedure: Maximum carbohydrate binding activity of the coupled lectins is retained. Linkage is stable over a range of pH values. Conjugated proteins are not leached off the beads by Tris or other routinely used buffers. No residual charges are present after conjugation. This minimizes non-specific binding to the matrix. Our agarose bound lectins are supplied at a constant concentration of lectin per ml of settled beads. The concentration for each lectin is selected to achieve the highest glycoconjugate binding capacity per mg of lectin present in the beads. Each lot is tested for its binding capacity using glycoproteins known to bind the lectin. This provides a guideline for the user and assures the quality of our agarose bound lectins. Inhibiting/Eluting Sugar: 200 mM N-acetylgalactosamine. *4 mg lectin/ml gel.

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