Surface-enhanced Raman scattering (SERS) spectroscopy has attracted considerable attention recently as a powerful detection platform in biosensing because of the wealth of inherent information ascertained about the chemical and molecular composition of a sample. However, real-world samples are often composed of many components, which renders the detection of constitutes of mixed samples very challenging for SERS sensing. Accordingly, separation techniques are needed before SERS measurements. Thin layer chromatography (TLC) is a simple, fast and costeffective technique for analyte separation and can a play pivotal role for on-site sensing. However, combining TLC with SERS is only successful to detect a limited number of analytes that have large Raman scattering cross sections. As a kind of biogenic amine, histamine (2-(4-imidazolyl)-ethylamine) has a relationship with many health problems resulting from seafood consumption occurring worldwide. Diatomaceous earth consists of fossilized remains of diatoms, a type of hard-shelled algae. As a kind of natural photonic biosilica from geological deposits, it has a variety of unique properties including highly porous structure, excellent adsorption capacity, and low cost. In addition, the two dimensional periodic pores on diatomite earth with hierarchical nanoscale photonic crystal features can enhance the localized optical field. Herein, we fabricate TLC plates from diatomite as the stationary phase combining with SERS to separate and detect histamine from seafood samples. We have proved that the diatomite on the TLC plate not only functions as stationary phase, but also provides additional Raman enhancement, in which the detection limit of 2 ppm was achieved for pyrene in mixture.