title: Application of screen-printed microband biosensors to end-point measurements of glucose and cell numbers in HepG2 cell culture.
abstract: Microband glucose biosensors were produced by insulating and sectioning through a screen-printed, water-based carbon electrode containing cobalt phthalocyanine redox mediator and glucose oxidase enzyme. Under quiescent conditions at 37 degrees C, at an operating potential of +0.4V, they produced an amperometric response to glucose in buffer solutions with a sensitivity of 26.4 nA/mM and a linear range of 0.45 to 9.0 mM. An optimal pH value of 8.5 was obtained under these conditions, and a value for activation energy of 40.55 kJ mol(-1) was calculated. In culture medium (pH 7.3), a sensitivity of 13 nA/mM was obtained and the response was linear up to 5 mM with a detection limit of 0.5 mM. The working concentration was up to 20 mM glucose with a precision of 11.3% for replicate biosensors (n=4). The microband biosensors were applied to determine end-point glucose concentrations in culture medium by monitoring steady-state current responses 400 s after transfer of the biosensors into different sample solutions. In conjunction with cultures of HepG2 (human Caucasian hepatocyte carcinoma) cells, current responses obtained in 24-h supernatants showed an inverse correlation (R(2)=0.98) with cell number, indicating that the biosensors were applicable for monitoring glucose metabolism by cells and of quantifying cell number. Glucose concentrations determined using the biosensor assay were in good agreement, for concentrations up to 20mM, with those determined spectrophotometrically (R(2)=0.99). This method of end-point glucose determination was used to provide an estimated rate of glucose uptake for HepG2 cells of 7.9 nmol/(10(6) cells min) based on a 24-h period in culture.
authors.affiliation: Centre for Analytical, Materials, and Sensors Science, Faculty of Health and Life Sciences, University of the West of England, Bristol, UK.
authors.forename: R M
authors.initials: RM
authors.lastname: Pemberton
authors.forename: J
authors.initials: J
authors.lastname: Xu
authors.forename: R
authors.initials: R
authors.lastname: Pittson
authors.forename: N
authors.initials: N
authors.lastname: Biddle
authors.forename: G A
authors.initials: GA
authors.lastname: Drago
authors.forename: S K
authors.initials: SK
authors.lastname: Jackson
authors.forename: J P
authors.initials: JP
authors.lastname: Hart
mesh_terms: D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D002452:Cell Count / Q000295:instrumentation*; D045744:Cell Line, Tumor; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods; D005947:Glucose / Q000032:analysis*; D006801:Humans; D015203:Reproducibility of Results
chemical_list: D005947:Glucose