The abuse of fentanyl and fentanyl analogs is becoming a global problem, with over 70,000 deaths reported in the USA, and there is evidence of its abuse beginning to appear in Europe and other areas. Fentanyl is 100 times more potent than morphine, and there are fentanyl analogs that are even more dangerous. Therefore, detecting and differentiating fentanyl isomers is critical in tracking this epidemic of drug use. Surface-enhanced Raman spectroscopy (SERS) is a useful method for detecting fentanyl isomers due to its capability of yielding spectroscopic fingerprints of the analytes. In this study, theoretical and experimental methods were compared in order to distinguish para fluorofentanyl, which is one of the most common fentanyl analogues in illicit mixtures, and its isomers ortho fluorofentanyl and meta fluorofentanyl. First, density functional theory (DFT) calculations were performed to define which vibrational peaks can be utilized in the identity of these analogs. The spectra obtained from the theoretical calculations were then compared with the spectra obtained from the Normal Raman and SERS experiments. To enhance sensitivity, bimetallic gold/silver nanostars (Au/Ag NS) were synthesized to provide the SERS enhancement along with magnesium chloride or potassium bromide aggregating agents with limits of detection in the low ng/mL range. The LOD value for para-fluorofentanyl is 3 ng/mL. The obtained results show that SERS is a successful technique to detect isomers of fentanyl analogs.