The novel immobilization technique using a poly(amino acid) multilayer was designed for the SPR sensing platform. These poly(amino acid) multilayer was confirmed as a relatively hydrophilic surface with little tendency for nonspecific adsorption of biomolecules and a flat surface that the biomolecular interactions can occur uniformly on the sensing surface. The poly(amino acid) multilayer functionalized with biotin was investigated to control the surface density and to estimate the optimum space between ligands for effective SPR sensing. This multilayer study demonstrates the importance of surface density which is significantly sensitive for fabrication of surface functional group in the monitoring of kinetic phenomena. (c) 2006 Elsevier B.V. All rights reserved.