title: Amperometric microcells for alkaline phosphatase assay.
abstract: To develop simple electrochemical immunoassays, a screen printed amperometric microcell with graphite working and Ag/AgCl reference electrodes was tested for the determination of alkaline phosphatase enzyme (ALP) and anti-humanIgG conjugated ALP (alpha-hIgG-ALP) activity in 5-10 microl samples. To ensure reproducible, steady state conditions, the working electrode surface was coated with mass-transport controlling hydrogel layer. The kinetic response curves of the hydrogel coated electrodes were linear. In addition, the hydrogel layer reduced the nonspecific adsorption of the alpha-hIgG-ALP conjugate on the working electrode surface. The measurements were made in the range of 2 divided by 4000 mU ml(-1) enzyme activities using ascorbic acid 2-phosphate (AAP) as the enzyme substrate. AAP is commercially available, non-toxic and has excellent stability. The sensitivity of the determinations was about 71% of the sensitivity which could be achieved using p-aminophenylphosphate (PAPP), a not easily accessible and unstable enzyme substrate. The experimentally determined kinetic parameters of the ALP enzyme catalyzed reactions were the same with the bare and hydrogel layer coated electrodes.
authors.affiliation: Joint Graduate Program in Biomedical Engineering, The University of Memphis, University of Tennessee Health Science Center, 38152, USA.
authors.forename: Róbert E
authors.initials: RE
authors.lastname: Gyurcsányi
authors.forename: Andrea
authors.initials: A
authors.lastname: Bereczki
authors.forename: Géza
authors.initials: G
authors.lastname: Nagy
authors.forename: Michael R
authors.initials: MR
authors.lastname: Neuman
authors.forename: Ernö
authors.initials: E
authors.lastname: Lindner
mesh_terms: D000469:Alkaline Phosphatase / Q000097:blood*; D015415:Biomarkers / Q000097:blood; D015374:Biosensing Techniques / Q000379:methods*; D004796:Clinical Enzyme Tests / Q000379:methods*; D004563:Electrochemistry; D006801:Humans
chemical_list: D015415:Biomarkers; D000469:Alkaline Phosphatase