The invention discloses a protein stabilizer and a preparation method thereof. The protein stabilizer comprises the following raw materials: high molecular weight hyaluronic acid, medium molecular weight hyaluronic acid, low molecular weight hyaluronic acid, oligomeric hyaluronic acid, acetylated hyaluronic acid, sodium polyacrylate, disodium ethylene diamine tetraacetate dihydrate, phenylmethylsulfonyl fluoride, magnesium chloride, polyethylene glycol monomethyl ether, sodium chloride or potassium chloride, ProCin 950, ProCin 300, bovine serum albumin, collagen, dipropylene glycol or glycerol, and a phosphate or Tris-HCl buffer solution. The quality guarantee period and stability of various proteins can be greatly prolonged by using a compounding technology of large and small molecular hyaluronic acid, gelatin and the like and a synergistic effect of macromolecules, preservatives and the like, the proteins, such as polystyrene boards, latex beads, magnetic particles, nitrocellulose membranes and the like, on various surfaces or in solutions can be greatly prolonged, and the stabilizers also reduce the background by eliminating non-specific binding of sample matrices and other assay ingredients.