Acetic anhydride, dextran and monomethoxypolyethylene glycol and different modification methods were used for modification of L-asparaginase to maintain enzyme activity and completely remove its antigenicity. The results showed that the macromolecular modifiers PEG and dextran were better than the small molecular modifier acetic anhydride. For maintenance of enzyme activity and removal of antigenicity modification in the presence of substrate was better than absence of substrate and activated PEG2 was better than activated PEG1. When PEG2-L-asparaginase was modified in the presence of substrate, its antigenicity was completely removed, while more than 30% of native enzyme activity were still retained.