The normal modes of vibration of crystalline 2-methyl-4-nitroaniline (MNA) deposited on nanostructured poly(tetrafluoroethylene) (PTFE) are investigated by means of polarized FT-IR Spectroscopy. As shown in a previous publication, the interface between the PTFE substrate and the MNA overlayer presents two distinct structures produced by an appropriate control of the experimental growth conditions. In both cases, using a polarized beam oriented parallel or perpendicular to the PTFE sliding direction, we measured the dichroic ratios of the absorption peaks. The MNA molecule is also studied by means of ab initio quantum-mechanical methods to characterize the geometry and the molecular normal modes of vibrations. On the basis of the structure of the MNA/PTFE interface and the ab initio results, the experimental spectra are thoroughly discussed, and the assignment of the most intense absorption peaks are proposed.