On-line determination of glucose concentration throughout animal cell cultures based on chemiluminescent detection of hydrogen peroxide coupled with flow-injection analysis.
A flow-injection analysis (FIA) system for the on-line determination of glucose in animal cell cultures is described. The system is based on immobilized glucose oxidase (GOD). The hydrogen peroxide generated in the enzyme reaction is determined via a highly sensitive chemiluminescent reaction with luminol. Based on the measurement of the maximum emitted light intensity, the system was able to analyse hydrogen peroxide over the concentration range of 10(-7) to 10(-2) M. For glucose determination, the system has a linear range of 10(-5) to 5 x 10(-2) M glucose, with an r.s.d. of 3% at the 1 mM level (5 measurements). The influence of luminol and buffer concentrations, pH and temperature on the chemiluminescent reaction were investigated. The enzyme reactor used was stable for more than 4 weeks in continuous operation, and it was possible to analyse up to 20 samples per h. The system has been successfully applied to on-line monitoring of glucose concentration during an animal cell culture, designed for the production of human antithrombin III factor. Results obtained with the FIA system were compared with off-line results, obtained with a Yellow Springs Instrument Company Model 27 (YSI).
Department of Enzyme Technology, Gesellschaft für Biotechnologische Forschung GmbH, Braunschweig, F.R.G.
Y L
YL
Huang
S Y
SY
Li
B A
BA
Dremel
U
U
Bilitewski
R D
RD
Schmid
D000818:Animals; D002138:Calibration; D002478:Cells, Cultured; D005947:Glucose / Q000032:analysis*; D006861:Hydrogen Peroxide / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D008163:Luminescent Measurements
D006861:Hydrogen Peroxide; D005947:Glucose