title: Automated amplified flow immunoassay for cocaine.
abstract: An amplified flow immunoassay (AFIA) was developed for cocaine, which combines a noncompetitive immunoenzymometric assay (IEMA) with an on-line detection of the enzyme label alkaline phosphatase (ALP) by a substrate-recycling biosensor. In the IEMA, the analyte cocaine first binds to a labeled polyclonal anti-cocaine antibody. Then, the excess labeled antibody is separated on an affinity column that contains a perfusion chromatography carrier modified by immobilized cocaine. The unbound complexes of the analyte cocaine with the ALP-labeled antibody are detected postcolumn. The detector senses phenol produced by ALP from phenyl phosphate. As detector, an amperometric substrate-recycling biosensor was used, which consists of a Clark-type oxygen electrode covered by tyrosinase and pyrroloquinoline quinone-dependent glucose dehydrogenase. The lower limit of detection is 380 pM (38 fmol) for cocaine. The sampling rate is 26/h. Cocaine could be detected from "real samples" with an imprecision of +/- 10% (n = 3) and with a recovery of 49 +/- 3% for various concentrations. AFIA is generally important as a new approach for the fast detection of picomolar concentrations of haptens.
authors.affiliation: Institute of Biochemistry and Molecular Physiology, University of Potsdam, Luckenwalde, Germany.
authors.forename: C G
authors.initials: CG
authors.lastname: Bauer
authors.forename: A V
authors.initials: AV
authors.lastname: Eremenko
authors.forename: A
authors.initials: A
authors.lastname: Kühn
authors.forename: K
authors.initials: K
authors.lastname: Kürzinger
authors.forename: A
authors.initials: A
authors.lastname: Makower
authors.forename: F W
authors.initials: FW
authors.lastname: Scheller
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis; D000906:Antibodies; D001331:Automation; D015374:Biosensing Techniques; D003042:Cocaine / Q000032:analysis*; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods*; D010636:Phenols / Q000032:analysis
chemical_list: D000906:Antibodies; D010636:Phenols; D000469:Alkaline Phosphatase; D003042:Cocaine