High-performance liquid chromatography of proteins on deformed nonporous agarose beads: immuno-affinity chromatography, exemplified with human growth hormone as ligand and a combination of ethylene glycol and salt for desorption of the antibodies

Abstract

An affinity chromatography column packed with nonporous agarose beads derivatized with human growth hormone via carbonyldiimidazol was used for the purification of antibodies against human growth hormone from antiserum. Desorption with 1 M sodium chloride in 60% ethylene glycol at pH 9.8 gave 100% total recovery of the antibodies, as measured by radioimmunoassay. The adsorption/desorption process is discussed in terms of hydrophobic and electrostatic interaction (these interactions may be involved in the bond between antibody and antigen in a cooperative fashion). The binding capacity of the column was estimated at about 50 micrograms of antibodies per gram sedimented agarose beads.