Methods for making saccharide-protein glycoconjugates

A polysaccharide antigen conjugate prepared by a process comprising the steps of:

• (a1) reacting a carbonyl group at a reducing terminus of a polysaccharide with Y1 of an additional linker of Formula 1,
• wherein Formula 1 is Y1-L-Y2,
• wherein L is a linking moiety, Y1 and Y2 are each —NHNH2 groups;
• (a2) reacting Y2 of the additional linker with a linker, to form a polysaccharide-additional linker-linker compound in which the free terminus of the linker is an ester group;
• and
• (b) reacting the ester group of the linker with a primary amine group in a carrier molecule, to form a polysaccharide-additional linker-linker-carrier molecule conjugate in which the linker is coupled to the carrier molecule via an amide linkage.

A process for preparing a conjugate of a polysaccharide and a carrier molecule comprising the steps of:

• (a1) reacting a carbonyl group at a reducing terminus of the polysaccharide with Y1 of an additional linker of Formula 1,
• wherein Formula 1 is Y1-L-Y2,
• wherein L is a linking moeity, Y1 and Y2 are each —NHNH2 groups;
• (a2) reacting Y2 with the linker SIDEA, to form a polysaccharide-additional linker-linker compound in which the free terminus of the linker is an ester group;
• and
• (b) reacting the ester group of the linker with a primary amine group in a carrier molecule, to form a polysaccharide-additional linker-linker-carrier molecule conjugate in which the linker is coupled to the carrier molecule via an amide linkage.