Chitosan, a natural product, is inherently biodegradable, biocompatible, and nontoxic. These properties make chitosan ideal for inclusion in matrices designed for use in enzyme immobilization for clinical analysis. This study demonstrates the feasibility of using chitosan in electrochemical biosensor fabrication. The enzyme sulfite oxidase (SOX) was covalently immobilized onto the matrix of chitosan-poly(hydroxyethyl methacrylate) (chitosan-pHEMA), a natural/synthetic polymer hybrid obtainable via UV curing, p-Benzoquinone, which served as an electron transfer mediator, was coupled onto the polymer network for activation of the chitosan-pHEMA copolymer, after completion of the photo-induced polymerization reaction. The biological activity of the immobilized SOX and the electroactivity of the coupled p-benzoquinone were investigated.