Preparation of nanogel-immobilized porous gel beads for affinity separation of proteins: Fusion of nano and micro gel materials

We describe the preparation and evaluation of nanogel-immobilized porous gel beads (GB) for application as a protein purification medium. Nanogel particles (NP) that bind with the Fc fragment of immunoglobulin G (IgG) were immobilized on the pore surface of macroporous hard GB containing quaternary ammonium cations on the surface via multipoint electrostatic interactions. The amount of NPs that were irreversibly immobilized in 1 ml of GB slurry was determined to be ∼30 mg using fluorescent-labeled NPs. Images obtained via scanning electron microscopy established that the NPs were uniformly immobilized on the surface of the pores without blocking the macropores. The model target protein (IgG) was reversibly captured by the NP-immobilized GBs through NP-IgG interactions. NP-immobilized GBs have potential applications as novel affinity purification media for proteins, combining inexpensive and stable ligands with high-performance supports.