For the purpose of investigating the chemical enhancement of amorphous semiconductors as well as increasing the sensitivity of the surface-enhanced Raman spectroscopy (SERS) substrate, titanium dioxide (TiO 2 ) precursors were calcined at different temperatures to generate crystallized TiO 2 (c-TiO 2 ) and amorphous TiO 2 (a-TiO 2 ) nanosheets, respectively. Afterward, a two-dimensional (2D) a-TiO 2 /Ag nanosheet SERS substrate was successfully fabricated using electrostatic interaction between a-TiO 2 and Ag nanoparticles. In order to demonstrate a greater SERS sensitivity on a-TiO 2 /Ag compared to either c-TiO 2 or Ag nanoparticles alone, the SERS probe molecules rhodamine 6G (R6G) and malachite green (MG) were utilized. Based on the results of SERS detections for probe molecules and contaminants, it demonstrates that a-TiO 2 /Ag nanosheets produce highly sensitive and repeatable Raman signals. The detectable concentration limits for R6G and MG were found to be 10 −11 M and 10 –10 M, respectively. And it has been determined that the system exhibits an enhancement factor (EF) of up to 1 × 10 8 . The limit of detection for 4-mercaptobenzoic acid and alizarin red can both reach 1 × 10 –8 . Furthermore, a finite-difference time-domain simulation is performed in order to evaluate the magnetic field strength generated by Ag nanoparticles. As a result of the simulation, it is evident that the actual EF is smaller than the calculated one, leading support to the view that a-TiO 2 nanosheets have a beneficial effect on the chemical enhancement of SERS.