Abstract: A sandwich amperometric enzyme immunoassay with flow injection for α-fetoprotein in human serum has been developed with alkaline phosphatase as the enzyme label. p-Hydroxyphenyl phosphate was used as the substrate for alkaline phosphatase. The hydrolysis product, hydroquinone, was detected by oxidative amperometry in a flow injection system. The amperometric wall jet detector was fitted with a glassy carbon working electrode held at 350 mV vs. Ag/AgCl. The detection limit of hydroquinone in 30 mM borate buffer pH 9.5 was 1.2 x 10^-10 M (linearity range: 10^-9 - 5.12 x 10^-6 M. A detection limit for free alkaline phosphatase of 1.2 x 10^-15 M (linearity range: 10^-15 - 10^-13 M), or about 36 000 molecules, was observed (same borate buffer and incubations of 10 min at 25°C). These conditions were maintained for the amperometric α-fetoprotein immunoassay. For comparison purposes, a photometric detection system was set up, with p-nitrophenyl phosphate as enzyme substrate and the same pair of antibodies and incubation conditions. The detection limit for α-fetoprotein obtained by amperometry, 0.07 ng/ml (linearity range = 5-500 ng/ml), was 14 times lower than by photometry. The amperometric enzyme immunoassay correlates well with a commercial colorimetric immunoassay (r = 0.986, slope = 0.967, n = 240).

Abstract: We developed a new urea sensor for the online monitoring of hemodialysis adequacy. The biosensor consisted of an immobilized urease cartridge placed between magneto-inductive conductivity cells. The biosensor output was taken as the conductivity difference between these cells. The device was placed on the ultrafiltrate line of a paired filtration dialysis system. The amount of urease present in the cartridge was sufficient for the complete conversion to ammonium carbonate of urea up to 35 mmol/L. Agreement was good between the urea concentration by the biosensor method and an automated analyzer for seven patients: range 8.07-30.3 mmol/L (22.6-84.8 mg/dL blood urea nitrogen, BUN); intercept 0.20±0.1 mmol/L (0.55±0.4 mg/dL, BUN); slope 1.01±0.01; r 0.997; S-y/x 0.40 mmol/L (1.11 mg/dL BUN). The device proposed meets the requirements of accuracy, cost, ruggedness, and ease of use (no calibration required) for a biosensor to be used for continuous monitoring of hemodialysis.