Affinity cryogels were synthesized using tris(hydroxymethyl)aminomethane (Tris) as ligand for specific interactions with the lysozyme (LYZ). The cryogel was produced by cryocopolymerization at −12 °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 (149.5 mg/g) was achieved when 376 mg/mL of Tris and 3.06 mol/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.