Abstract：Magnetic γ-Fe2O3@Au core/shell nanoparticles were prepared by the growth of Au shell onto the seeds of γ-Fe2O3 nanoparticles. The γ-Fe2O3@Au nanoparticles were functionalized by the modification of antibody. By the specific interaction between antigen and antibody, one of the antigens was separated in order or two antigens were separated together with the assistance of external magnetic field. The efficiency of magnetic separation was evaluated based on surface enhanced Raman spectroscopy (SERS) immunoassay technique. The concentration limitation for the magnetic separation and the SERS evaluation were obtained. The results showed that the method of magnetic immune separation has been achieved to separate any components in the media with three antigens without any influence on the existence of other antigens. The concentration of the separated antigen was reduced to the detection limitation of SERS-based immunoassay which could reach about 0.1 pg/mL.