Protein immobilization on silica supports : A ligand density study

Abstract

The immobilization of proteins on diol-bonded silica matrices containing carboxyl groups (spacer arms) was studied. It was found that the activated ester coupling method worked best with N,N′-dicyclohexylcarbodiimide as the condensing agent in the activation step. During protein coupling, the amount of protein immobilized was highest below pH 6. The optimum pore size of the silica was 300–1000 Å. The spacer arm ligand density was varied over as much as a 100-fold range and the effects on the total activities and specific activities of several proteins were studied. Two proteins exhibited up to two-fold increases in specific activity at low ligand densities. However, the total amounts of activity and protein immobilized decreased at low ligand densities.