Abstract: Optical properties of histamine and l-histidine have been analyzed by using surface-enhanced Raman scattering (SERS). A silver film over nanosphere (AgFON) structure with 120-nm-thick silver film on polystyrene nanospheres 1,000 nm in diameter is fabricated by nanosphere lithography to enhance the Raman signal excited at the laser wavelength of 532 nm. Normal Raman spectrum and the SERS spectrum of histamine and l-histidine were compared. Further, vibration modes of these molecules were calculated by using density functional method. In the SERS experiment, we were able to measure the Raman spectrum with a histamine concentration as less as 100 pM. This sensitivity is higher than that from high-performance liquid chromatography.