The adsorption of ferritin from phosphate-buffered saline (PBS) onto gold has been examined using a quartz crystal microbalance (QCM), surface plasmon resonance (SPR), X-ray photoelectron spectroscopy (XPS), and atomic force microscopy (AFM). Application of these techniques allows elucidation of the surface coverage and layer thickness of ferritin on gold. The kinetics of ferritin adsorption onto gold were investigated by monitoring the resonant frequency shift of a QCM. Adsorption isotherms for ferritin in PBS and in air have been measured by QCM. These isotherms suggest less than monolayer coverage of ferritin on gold. A layer thickness of 4.8 +/- 0.2 nm was calculated for the dry ferritin film from QCM data. Measurements of the shift in the reflectivity of light at a fixed angle close to the SPR plasmon resonance were also used to follow the kinetics of ferritin adsorption. Full SPR curves were measured in PBS solution and air and were used to determine the effective thickness of the ferritin layer in both environments. The ferritin layer on gold from SPR data was found to be twice as thick when measured in PBS as it was for a dry film. This difference in thickness is attributed to shrinkage of ferritin with drying. Angle-resolved XPS measurements on a dry ferritin film (preadsorbed on gold) yield a ferritin thickness of 4.7 +/- 0.5 nm, a value in good agreement with those determined from QCM and SPR. QCM, SPR, and XPS all yield a surface coverage of 6.3 +/- 0.7 mg m(-2) for dry ferritin layers on gold. AFM enabled examination of the topography of ferritin adsorbed on gold on the nanometer scale and confirmed that ferritin forms an incomplete monolayer. In all cases, ferritin was found to be irreversibly adsorbed to gold and to form a stable protein layer, thus making it well suited as a biological receptor layer for immunosensing applications.