It is very important to prepare high sensitive surface enhanced Raman scattering (SERS) substrates in the SERS detection process. The preparation of metal nanoparticle aggregates by light manipulation technology is a hot topic in the field of SERS. In this paper, femtosecond laser wet etching technology was used to etch slot array with the cross-sectional area (width×depth) of 10 μm×7 μm, 30 μm×12 μm, 60 μm×15 μm, 70 μm×19 μm, 90 μm×21 μm in the range of 5 mm×5 mm on the surface of silicon wafer, a Silicon-based micro-nano structure substrate (SiMS) with the different cross-sectional area was prepared. SERS enhancement of analytes on the substrates was achieved using optical manipulation techniques combined with SERS technique. The laser was focused on the substrate slot, due to the action of the light radiation pressure, the gold nanoparticles move along the direction of the beam propagation and accumulate in the slots on the surface of the structure to form gold nanoparticle aggregates, which promote “hot spots” effect. The sensitivity of the SERS detection was improved, and the SERS enhancement of the probe on the substrates was achieved. Experiments show that the metal nanoparticles can effectively accumulate in the slot on the surface of the SiMS when the optical radiation pressure is greater than the optical gradient force, forming more “hot spots”, which can greatly improve the SERS enhancement effect. The SERS signal of pyrene is gradually enhanced with the increase of the cross-sectional area of the slot, the enhancement effect of the slot with width and depth of 70×19 μm2 was the best, SERS intensity of pyrene was increased by about two magnitudes, and the minimum detection concentration was 5.0×10-9 mol·L-1. Beyond this cross-sectional area, the SERS intensity begins to decrease. The lowest detection concentration of pyrene is 5.0×10-9 mol·L-1. In the low concentration range (5.0×10-9～1.0×10-7 mol·L-1). It demonstrated a good linear correlation between the SERS intensity of characteristic peaks at 588 and 1 234 cm-1 and concentration, and the fitting equation and linear correlation coefficient were 0.992 and 0.971, respectively. The SiMS with a cross-sectional area of 70×19 μm2 was used for repetitive experiments, and eight different positions on the substrate were selected. After each position was measured, the laser was switched off and the action of the laser was disappeared, the gold nanoparticles were re-dispersed in the solution. Three measures were repeated for each position. The relative standard deviation (RSD) of the two peaks at 588 and 1 234 cm-1 at different positions of the substrate were 9.9% and 2.0%, respectively, which showed good repeatability. The study showed that the SERS effect could be greatly improved by the optical manipulation-SERS method based on the SiMS and this method has potential for application in the detection and analysis of materials in fields such as chemistry and biology.