THE USE OF CULTURE REDOX POTENTIAL AND OXYGEN-UPTAKE RATE FOR ASSESSING GLUCOSE AND GLUTAMINE DEPLETION IN HYBRIDOMA CULTURES

Culture redox potential (CRP) and oxygen uptake rate (OUR) were monito red on-line during glucose- and glutamine-limited batch cultures of a murine hybridoma cell line that secretes a neutralizing monoclonal ant ibody specific to toxin 2 of the scorpion Centruroides noxius Hoffmann . It was found that OUR and CRP can be used for assessing the viable c ell concentration and growth phases of the culture. Before nutrient de pletion, OUR increased exponentially with viable cell concentration, w hereas CRP decreased monotonically until cell viability started to dec rease. During the death phase, CRP gradually increased. A sudden decre ase in OUR occurred upon glucose or glutamine depletion. CRP traced th e dissolved oxygen profile during a control action or an operational e ventuality, however, during nutrient depletion it did not follow the e xpected behavior of a system composed mainly by the O-2/H2O redox coup le. Such a behavior was not due to the accumulated lactate or ammonia, nor to possible intracellular redox potential changes caused by nutri ent depletion, as inferred from respiration inhibition by rotenone or uncoupled respiration by 2,4-dinitrophenol. As shown in this study, op erational eventualities can be erroneously interpreted as changes in O UR when using algorithms based solely on oxygen balances. However, sim ultaneous measurements of CRP and OUR may be used to discriminate real metabolic events from operational failures. The results presented her e can be used in advanced real-time algorithms for controling glucose and glutamine at low concentrations, avoiding under-or over-feeding th em in hybridoma cultures, and consequently reducing the accumulation o f metabolic wastes and improving monoclonal antibody production. (C) 1 997 John Wiley & Sons, Inc.