title: An electrochemiluminescence-based fibre optic biosensor for choline flow injection analysis.
abstract: A fibre optic biosensor based on luminol electrochemiluminescence (ECL) integrated in a flow injection analysis (FIA) system was developed for the detection of choline. The electrochemiluminescence of luminol was generated by a glassy carbon electrode polarised at +425 mV vs. a platinum pseudo-reference electrode. Choline oxidase (Chx) was immobilised either covalently on polyamide (ABC type) or on UltraBind preactivated membranes, or by physical entrapment in a photo-cross-linkable poly(vinyl alcohol) polymer (PVA-SbQ) alone or after absorption on a weak anion exchanger, DEAE (diethylaminoethyl) Sepharose. The optimisation of the reaction conditions and physicochemical parameters influencing the FIA biosensor response demonstrated that the choline biosensor exhibited the best performances in a 30 mM veronal buffer containing 30 mM KCl and 1.5 mM MgCl2, at pH 9. The use of a 0.5 ml min-1 flow rate enabled the measurement of choline by the membrane-based ECL biosensors in 8 or 5 min, with ABC or UltraBind membranes, respectively, whereas the measurement required only 3 min with the DEAE-PVA system. For comparison, the detection of choline was performed with Chx immobilised using the four different supports. The best performances were obtained with the DEAE-PVA-Chx sensing layer, which allowed a detection limit of 10 pmol, whereas with the ABC, the UltraBind and the PVA systems, the detection limits were 300 pmol, 75 pmol and 220 pmol, respectively. The DEAE-based system also exhibited a good operational stability since 160 repeated measurements of 3 nmol of choline could be performed with an RSD of 4.5% whereas the stability under the best conditions was 45 assays with the other supports.
authors.affiliation: Laboratoire de Génie Enzymatique, UMR CNRS 5013, Université Claude Bernard Lyon 1, Villeurbanne, France.
authors.forename: V C
authors.initials: VC
authors.lastname: Tsafack
authors.forename: C A
authors.initials: CA
authors.lastname: Marquette
authors.forename: B
authors.initials: B
authors.lastname: Leca
authors.forename: L J
authors.initials: LJ
authors.lastname: Blum
mesh_terms: D015374:Biosensing Techniques; D002794:Choline / Q000032:analysis*; D005336:Fiber Optic Technology; D017022:Flow Injection Analysis; D008163:Luminescent Measurements
chemical_list: D002794:Choline