Crystallization of hen egg white lysozyme (HEWL) from aqueous solution on behenic acid films was investigated by atomic force microscopy and optical microscopy. It was found that the tetragonal HEWL crystals prefer to nucleate with the {101} planes parallel to thick behenic acid films and bare mica substrates. In contrast, in addition to the {101} crystal, {101}-oriented crystals also nucleated on monomer layer films of behenic acid. The ratio of {110}-oriented crystals was strongly influenced by the coverage of the monomer layer films. The influences of the hydrophilicity of the substrates on the HEWL crystal nucleation were also discussed. (C) 2004 Elsevier B.V. All rights reserved.