A novel one-step protocol for the preparation of sub-micron magnetic particles as small as 30 nm in diameter has been developed. The surface of the par tides was functionalized with carboxyl groups (similar to 1 COOH per 15 Angstrom(2)) to facilitate the attachment of affinity ligands. The high surface area of the resulting beads, combined with the high density of functional groups on the surface, makes them ideally suited for the preparative purification of proteins as was demonstrated by the efficient isolation of trypsin (36 mg per gram of particles) from pancreatic extract. (C) 1998 John Wiley & Sons, Inc.