A label-free amperometric immunosensor for the detection of (x-fetoprotein (AFP) based on the layer-by-layer (LBL) assembly of positively charged multi-walled carbon nanotubes-chitosan (MCNTs-CS) and negatively charged platinum-Prussian blue hybrid nanoparticles (Pt-PB) on 3-mercaptopropanesulfonic, sodium salt (MPS) modified electrode is described in this paper. Transmission electron microscopy (TEM) was employed to characterize the morphologies of Pt-PB and Pt-PB-MCNTs-CS. The stepwise LBL assembly process of electroactive species on electrode surface was evaluated by cyclic voltammetry (CV). The immunosensor shows good analytical performance such as low detection limit (0.08 ng ml(-1)), wide linear range (from 0.1 to 15.0 and from 15.0 to 200.0 ng ml(-1)), good regeneration, selectivity, stability, and reproducibility for the determination of AFP. Application of the immunosensor to clinical samples demonstrated that results were in good agreement with ELISA. (C) 2008 Elsevier B.V. All rights reserved.