Binding layer and method for its preparation and uses thereof

A method for preparing a binding layer comprising a polysaccharide substituted by a plurality of carboxylic groups, the plurality of carboxylic groups comprising a plurality of easily-activated carboxylic groups, each of the plurality of the easily-activated carboxylic groups having at least one β-atom being a carbon atom and remaining β-atoms are independently hydrogen or carbon atoms, the method comprising:

• providing a substrate having a surface;
• immobilizing a polysaccharide on the surface of the substrate wherein said polysaccharide immobilized on the substrate contains functional groups capable of chemical modification,
• chemically modifying the polysaccharide by attaching a plurality of the easily-activated carboxylic groups to extend therefrom, wherein at least one β-atom of each group of the easily-activated carboxylic group is a carbon atom and remaining β-atoms are independently hydrogen or carbon atoms, wherein said attaching comprises linking the β-carbon atom of the plurality of the easily activated functional group to the functional groups of the polysaccharide through a covalent linkage, the functional groups being a native or chemically introduced carboxylic groups, or a derivative thereof selected from the group consisting of an ester, an amide, an anhydride, and an acetyl halide;
• and
• esterifying at least part of the easily activated carboxylic groups for binding of ligand molecules with at least one ester selected from the group consisting of a N-hydroxysuccinimide ester and a N-hydroxysulfosuccinimide ester,
• wherein chemically modifying and esterifying may occur, independently of each other, either before, during or after immobilizing, and a β-atom of the easily-activated carboxylic groups has no electron withdrawing groups.