The invention discloses a preparation method of a low-abundance low-molecular-weight protein-rich material. The preparation method comprises the following steps of: fixing a P-type boron-doped silicon wafer in an electrolytic cell; adding ethanol and hydrofluoric acid as electrolyte; performing direct current electrolytic etching and demolding by taking the silicon wafer as an anode and a platinum electrode as a cathode; cleaning an etched and demolded porous silicon layer with ethanol, then ultrasonically crushing and blowing with nitrogen; performing ozone oxidation treatment on porous silicon particles; soaking the oxidized porous silicon particles in mixed solution of silylation reagent and organic solvent to react, then drying in a drying box, and soaking and cleaning the porous silicon particles, which are dried by evaporation, with ethanol; and selecting porous silicon with required aperture and performing surface modification by using amino silylation reagent, sulfydryl silylation reagent or epoxy silylation reagent. According to the preparation method, the effect of separating, enriching and detecting low-molecular-weight protein in a blood serum sample in one step is achieved by preparing the porous silicon particles.