Affinity cryogels were synthesized using tris(hydroxymethyl)aminomethane (Tris) as ligand for specific interactions with the lysozyme (LYZ). The cryogel was produced by cryo-copolymerization at -12 degrees C. A central composite rotational design 2(2) was used to optimize the immobilization procedure of the Tris on the cryogel. A maximum adsorption for LYZ (149.5mg/g) was achieved when 376mg/mL of Tris and 3.06mol/L of sodium borohydride were used during the Tris immobilization. Chromatographic separation of LYZ from chicken egg white was done with a purity of 92.13%. Results showed that the affinity cryogel was a potential separation medium for LYZ purification.