Oppositely charged polyelectrolyte assemblies of poly(acrylic acid) and poly-L-lysine are deposited as alternate layers on cystamine-functionalized Au surfaces. Microgravimetric, quartz-crystal-microbalance, measurements and ellipsometric studies reveal a nonlinear increase in the polymer assembly thickness upon the buildup of the polymer layers. This is attributed to the swelling of the polymer upon the stepwise assembly of the layered film. The interfacial and intra-assembly properties of the polyelectrolyte systems were characterized by Faradaic impedance spectroscopy. In the presence of Fe(CN)(6)(3-)/Fe(CN)(6)(4-) as the redox label, all assemblies that terminate with a negatively charged interface are characterized by a high interfacial electron-transfer resistance, originating from the electrostatic repulsion of the redox label from the interface. The interfacial electron-transfer resistance also increases as the number of layers, or assembly thickness, increases. For assemblies terminated with a positively charged polyelectrolyte, a very low interfacial electron-transfer resistance for the redox process of the negatively charged redox probe is detected. This is attributed to a neutralized, porous structure of the polymer assembly. For the positively charged redox label, protonated N,N-dimethylaminomethyl-ferrocene, similar results are observed for the assemblies with the opposite dependence on the charge of the terminal layer. The electrodes functionalized with the polymers were used for the selective oxidation of a mixture consisting of Fe(CN)(6)(4-) and protonated N,N-dimethylaminomethyl-ferrocene.