A silica-based adsorbent for affinity chromatography of serine protease was prepared by bonding p-aminobenzamidine (pABZ) to aminopropyl silica. Silanization of silica with both gamma-aminopropyltrimethoxysilane and gamma-aminopropyltriethoxysilane under anhydrous conditions led to a monolayer density of primary amino groups. The aminopropyl silica was converted to primary hydroxyl-containing silica via diazotization, and consequently activated with p-nitrophenyl chloroformate and immobilized with ligand pABZ. The resultant silica-pABZ has a ligand density of 13.8 mumol g-1. Pyridine was indicated by the data to increase the reactivity of the chloroformate toward the hydroxyl group more efficiently than 4-dimethylaminopyridine. Two stages of adsorption were found in batch adsorption of trypsin with an equilibrium adsorption isotherm of the Langmuir type. When the chromatographic column packed with this silica-pABZ was operated under a higher flow rate (2.33 cm3 min-1) and with 3 g dm-3 of crude urokinase as the influence, the yield was 55%. Both the flow rate and the concentration of the crude protein were shown, by measuring the dynamic binding capacity from chromatographic experiments, to be the factors which influenced the chromatographic efficiency.