The formation process and structure of a self-assembled monolayer (SAM) of lipoic-acid-terminated polyproline on a gold surface in aqueous solution were investigated by several techniques. The amount of polyproline molecules on the gold surface was determined from the area of the reductive desorption peak, and orientation and thickness of the polyproline SAM were determined in situ by attenuated total reflection infrared (ATR-IR) spectroscopy and ellipsometry. The kinetics of the polyproline SAM formation process were discussed on the basis of these results. The in situ IR study confirmed that the conformation of the polyproline SAM was changed by changing the solvent from water to methanol and methanol to water, as is the case for polyproline dissolved in solution.