Abstract: Xylanase was immobilized with gelatin and cross linked with glutaraldehyde. The effect of reaction conditions (the concentration of gelatin, glutaraldehyde concentration, cross-linking time and immobilized time) on the relative activity of immobilized xylanase were tested. The most optimal immobilized conditions were determined with orthogonal test. At the same time the most suitable immobilized temperature, thermal stability, the most appropriate reacted pH and the pH stability of the immobilized xylanase were compared with that of the free enzyme. The result showed that the best immobilized condition was obtained. The recovery rate of immobilized xylanase was 72.56% when the concentration of gelatin was 15%, the volume fraction of glutaraldehyde was 4%, the cross-linked time was 1h and the immobilized time was 3h. Simultaneously the most suitable immobilized temperature was respectively 50℃ and 60℃, the most appropriate pH was respectively 3.6 and 4.6 of the immobilized xylanase and the free enzyme. The pH stability and the thermal stability were obviously improved.