In the real world, analytes usually exist in complex systems, and this makes direct detection by surface-enhanced Raman spectroscopy (SERS) difficult. Thin layer chromatography tandem with SERS (TLC-SERS) has many advantages in analysis such as separation effect, instant speed, simple process, and low cost. Therefore, the TLC-SERS has great potential for detecting analytes in mixtures without sample pretreatment. The review demonstrates TLC-SERS applications in diverse analytical relevant topics such as environmental pollutants, illegal additives, pesticide residues, toxic ingredients, biological molecules, and chemical substances. Important properties such as stationary phase, separation efficiency, and sensitivity are discussed. In addition, future perspectives for improving the efficiency of TLC-SERS in real sample detecting are outlined.