Expanded poly(tetrafluoroethylene) (ePTFE) films grafted with 2-hydroxyethyl methacrylate (HEMA) and 2-hydroxyethyl acrylate (HEA) were applied to a polymer support for urease immobilization. The HEMA-and HEA-grafted ePTFE (ePTFE-g-PHEMA and ePTFE-g-PHEA) films prepared by the combined use of the plasma treatment and photografting possessed high water-absorptivities. Imidazole groups were introduced to grafted PHEMA and PHEA chains with 1,1'-carbonyldiimidazole (CDI) in acetonitrile. The activity of urease covalently immobilized to the ePTFE-g-PHEMA and ePTFE-g-PHEA films in a pH 7.0 buffer at 4 degrees C had the maximum value at the optimum pH value of 7.5 for native urease. Urease immobilized on the ePTFE-g-PHEMA films with the extent of CDI bonding of about 20% had the maximum activity, and the repeatedly measured activity was kept almost constant. The relative activity of immobilized urease stayed almost constant in the range of the immobilized amounts between 10 and 30 mg/g for both grafted ePTFE films, and decreased at higher immobilized amounts because of the crowding of immobilized urease molecules in the grafted layers. The relative activity of immobilized urease had the maximum values at the grafted amounts of 1.2 and 1.7 mmol/g for the ePTFE-g-PHEMA and ePTFE-g-PHEA films, respectively, and the further increase in the grafted amount resulted in the decrease in the relative activity. The optimum temperature of the activity for immobilized urease was shifted from 30 to 50 degrees C for native urease by the covalent immobilization on both grafted ePTFE films and immobilized urease was repeatedly usable without a considerable decrease in the activity in the regions of the pH 6.0-9.0 and 10-60 degrees. (c) 2006 Wiley Periodicals, Inc.