ONLINE MONITORING OF GLUCOSE IN MAMMALIAN-CELL CULTURE USING A FLOW-INJECTION ANALYSIS (FIA) MEDIATED BIOSENSOR

A flow injection analysis (FIA) biosensor system has been developed fo r on-line determination of glucose during mammalian cell cultivation. The culture sample was peristaltically withdrawn from the bioreactor a nd after cell separation by a steam sterilizable ceramic microfilter, the filtrate was continuously fed to the FIA mediated-biosensor system at 4 mLh(-1), whereas the cell-containing retentate was recirculated to the bioreactor. In the amperometric biosensor system, glucose oxida se was covalently immobilized onto a preactivated nylon membrane and a ttached to the sensing area of a platinum working electrode. The enzym e reaction was coupled with the mediator 1,1'-dimethylferricinium (DMF e+)-cyclodextrein inclusion complex to recycle the reduced glucose oxi dase to its original active state. 1,1'- dimethylferrocene (DMFe) was then reoxidized to DMFe+ at the surface of the platinum electrode pois ed at + 0.15 V vs silver/silver chloride. The FIA mediated-biosensor w as linear up to 6 mM glucose, with a detection limit of 0.1 mM, and po ssessed excellent reproducibility (+/-0.4%, 95% confidence interval) o ver 123 repeated analyses during a 62 h continuous operation. The immo bilized glucose oxidase was stable for up to 7 days when applied to gl ucose measurement during 5-10 day fed-batch cultivation of 293S mammal ian cells. The results obtained from the mediated-biosensor system com pared well with the hexokinase and HPLC data. (C) 1997 John Wiley & So ns.