The buildup of layer-by-layer assemblies onto gold surfaces from water-soluble charged polyelectrolytes and proteins is examined using quartz crystal microgravimetry (QCM) and electrochemical techniques. Polyelectrolytes such as poly(styrenesulfonate) and poly(ester sulfonic acid) (Eastman AQ-29D polymer) adsorb spontaneously onto gold, contrary to poly(ethyleneimine). From the modification of the gold surface with a thiol and specific adsorption of polymers under polarization conditions, it is concluded that the hydrophobicity of the gold surface seems to be a determining factor in the adsorption process. Alternate adsorption onto gold resonators first coated with AQ-29D polymer gives stable multilayer films in the case of positively charged lysozyme (pI = 11) or polyheme Desulfovibrio vulgaris Hildenborough cytochrome c(3) (PI = 10.5). QCM frequency changes with the number of adsorption steps suggest that a linear increase in film mass occurs. Desulfomicrobium norvegicum polyheme cytochrome c(3) (PI = 7), which has a null global charge at neutral pH, is shown to give also stable multilayer AQ-29D/cytochrome c(3) films, suggesting that several types of interactions, especially the hydrophobic effect, are involved in the buildup process.