Diamine oxidase was purified over 2300-fold from porcine kidney to a specific activity of 1 U mg(-1). The final preparation exhibited a single 103 kDa protein band and contained relatively large amounts of Asx and Glx acidic residues. When stored at -80 degrees C, soluble enzyme retained its original catalytic activity for at least five months. The optimal pH and temperature of the enzyme immobilized by intramolecular cross-linking via gill taraldehyde activation and deposited onto a preactivated nylon membrane were 7.4 and 60 degrees C with cadaverine as substrate. The apparent K-m(n) values (Michaelis-Menten constants) of immobilized diamine oxidase with histamine, putrescine, and cadaverine as substrates were estimated to be 0.27. 3.2, and 0.64 mM, respectively. Artificial mediators such as ferrocene derivatives, 2,6-dichlorophenolindophenol, potassium ferricyanide and 4-aminodiphenylamine were nor observed to facilitate electron transfer from the reduced enzyme to the electrode. The biosensor using the immobilized diamine oxidase and a platinum working electrode (poised at +700 mM vs Ag/AgCl for determination of hydrogen peroxide released from the enzymatic oxidation) was linear up to 6 mM histamine, cadaverine, or putrescine with a lower detection limit of 25 mu M. Each analysis could be performed in 3 min including washing and time for the current to return to baseline. The enzyme membranes were stable at 5 degrees C for at least two months and could be used for more than 60 repeated analyses without significant loss of sensitivity.