The invention discloses a preparation method and SERS application of monodispersed silver-shell magnetic microspheres. In a base, 400 nm Fe3O4 microspheres are taken as cores to provide magnetism, 50 nm SiO2 shells are taken as hydrophilic sandwich layers to improve the dispersibility of the microspheres, and complete and uniform silver shells are prepared with chemical plating and seed reduction methods. The preparation method comprises the following steps: enabling the 400 nm Fe3O4 microspheres to be covered with the 50 nm SiO2 shells with an improved Stober method, and then carrying out sensitization on the SiO2 shells through SnCl2, so that a great deal of Sn<2+> are adsorbed; washing the magnetic microspheres with deionized water, after the SnCl2 is completely removed, and using a 40 mM silver-ammonia solution to perform kernelization on Fe3O4@SiO2 under an ultrasonic condition, so that a large quantity of Ag seeds are generated in an in situ manner; finally using a seed growing method, and under high-concentration PVP protection, using formaldehyde and stronger ammonia water to reduce the complete and uniform Ag shells so as to form the monodispersed silver-shell magnetic microspheres. The invention also discloses application of the silver-shell magnetic microspheres prepared through the method in SERS detection, the monodispersed silver-shell magnetic microspheres are high in magnetic responsiveness, good in dispersibility, stable in structure and extremely high in enhancing effect, and can be directly used for SERS detection of various micromolecular pollutants, pesticide residues, serum, cancer biomarkers and the like under a solution system.