We report the construction of three-dimensional (3D) colloidal crystal arrays (CCA) and hollow colloidal crystal arrays (HCCA) derived from the self-assembly of polyelectrolytes (PE)-coated polystyrene (PS) particles and their use as models of high surface area systems to immobilize peroxidase (POD). POD molecules could infiltrate into the deep layers of CCA and HCCA through their interconnected pores and strongly adsorbed at the PE shell of the colloidal particles. And the total enzyme loading amount and bioactivities increased linearly with the thickness of the CCA till ca. 10 mu m. Compared with flat substrates with the same geometrical area, CCA and HCCA exhibit much higher enzyme loading abilities (similar to 43 and 53 times respectively) and the resulting bioactivities (similar to 35 and 41 times respectively) due to their inherently higher surface area and 3D interconnected porous structures. In addition, HCCA could load approximately 30% more POD than CCA because some POD molecules could infiltrate into the interior of the hollow capsule under salt condition. (C) 2008 Elsevier Inc. All rights reserved.