Several biosensing techniques like SPR and QCM use glass as the substrate, bearing a thin Au layer which serves as the sensing surface. Other substrates of interest include Si due to processing knowledge and the potential integration of electronics, and Cytop which has a refractive index close to that of water. Au on glass is highly crystalline, with a large 111 character, especially after annealing, whereas on Si and on Cytop, Au is non-textured ("polycrystalline"). These structural differences result in distinct voltammetric behavior, both faradaic and capacitive, of SAM- and protein-coated Au on different substrate types. A contact angle and double-layer capacitance examination of protein adsorption on SAM-coated Au on these substrates, and electrochemical desorption of the resulting molecular stacks, indicates that the usual assumption that hydrophobic SAMs promote adsorption and hydrophilic ones prevent adsorption is not universally true, and depends both on the protein type and the substrate. Thus, caution is advised against applying results obtained on one substrate to another. On the other hand, systems utilizing bioconjugation seem to be much less sensitive to Au type, and thus the underlying substrate. Electrochemical desorption of proteins from Au does occur but with limited efficiency. (C) 2012 The Electrochemical Society. [DOI: 10.1149/2.034301jes] All rights reserved.