Surface-enhanced Raman scattering (SERS) has been widely used in food and drug detection, biological and medical sensing. In recent years, the study of non-metallic SERS substrates has gradually become a hot field of SERS. Here, we investigate the modulation effect on SERS activities of 2,3,5,6-tetrafluoro-7,7,8,8-tetrachyanoquindimethylene (F ₄TCNQ) grown on molybdenum disulfide (MoS ₂) films. The different nanostructures of F ₄TCNQ can have an effect on the bound capability of charges transferred from the surface of MoS ₂, which changes the electron density distribution on the surface of the F ₄TCNQ/MoS ₂nanocomposite material. Therefore, the interface exhibits different charge localizations in the F ₄TCNQ/MoS ₂nanocomposite. The charge transfer efficiency between the substrate and the adsorbed probe molecules leads the substrate to show a different SERS sensitivity. The enhancement factor of 4-mercaptobenzoic acid (4-MBA) molecules on the most optimized 7-min F ₄TCNQ/MoS ₂nanocomposite substrate can reach $ 6.9\times {10}^{4} $ , and the detection limit concentration is as low as 10 ^–6mol/L. The result of research on F ₄TCNQ/MoS ₂nanocomposite provides an effective optimization scheme of energy level regulation for SERS based on the chemical enhancement mechanism, and opens up a new way to further exploit its functional applications.