Mammalian cell culture: engineering principles and scale-up

229

The negative effects of excessive agitation on tissue cells in microcarrier culture have often been ascribed to "shear." Analysis of the fluid mechanics occurring suggests that there are actually three potential damage mechanisms: collisions of a cell-covered microcarrier with other beads, collisions with parts of the reactor (primarily the impeller), and interaction with turbulent eddies the size of the microcarrier beads. Review of the available quantitative information on agitation effects in cell cultures does not establish which mechanism is predominant; the range of experimental variables reported emphasizes power input over the other reactor and impeller parameters. The bead-bead collision model is tentatively supported by the available data, but the other mechanisms may still be significant in some systems. The formation of bead aggregates by cellular bridging provides a parallel means of damaging cells. Breaking of these bridges by any of the three means identified earlier can cause cell destruction and/or the net transfer of cells to formerly bare beads. High concentrations of bridges are favored by lower agitation rates, presumably because the bridges are not as quickly destroyed after formation.

Section: Literature Datamentioning

confidence: 72%

Physical mechanisms of cell damage in microcarrier cell culture bioreactors

Cherry

Papoutsakis

1988

229

“…Although the contributions from glycolysis and respiration were different after the step change, q A T p was unchanged, which is consistent with results obtained in batch culture of MDCK cells using different sugars. 6 Lower by-product yields might be expected after the step change because less glutamine is available for biosynthetic reactions, but Y&, and YkH3,&ln are essentially unchanged (Fig. 6c).…”

Section: Steady-state Metabolic Changesmentioning

confidence: 93%

“…This explanation is also consistent with the increased rate of ATP synthesis observed during the lag phase of FS-4 cell microcarrier culture. 6 The responses of qATp and the cell concentration to the step change can be used to estimate Y,,Ap and the maintenance requirements of the cells. The qATp was above the baseline value of 23.3 X mmol ATP/cell day for about 36 h. During this time, the cells produced 12.7 mmol ATP/L in addition to the amount required to maintain a specific growth rate of 0.66 day-'.…”

Section: Cell Growth and Energeticsmentioning

confidence: 99%

Transient responses of hybridoma cells to nutrient additions in continuous culture: I. Glucose pulse and step changes

Miller

Wilke

Blanch

1989

105

Glucose and glutamine are the main nutrients used by mammalian cells in culture. Each provides unique biosynthetic precursors but are complementary for production of other metabolites and energy. The transient and steady-state responses of hybridoma growth and metabolism to glucose pulse and step changes have been examined. Metabolic quotients are reported for oxygen, glucose, lactate, ammonia, glutamine, alanine, and other amino acids. The glucose consumption rate increased by 100-200% immediately after glucose was added to the reactor, and the increased glycolytic ATP production appears to be responsible for the concurrent rapid decrease in the oxygen consumption rate. The effects on glutamine consumption were delayed, probably due to buffering by the TCA cycle and interrelated pathways. A period of increased biosynthetic activity, as evidenced by an increase in the estimated specific ATP production rate and lower by-product yields from glutamine, preceded the increase in cell concentration after the glucose step change. The biosynthetic yield of cells from ATP was calculated, and it was estimated that maintenance accounted for about 60% of the energy used by the cells at a specific growth rate of 0.66 day(-1). The estimated 22% ATP production due to glycoysis was twice as great as that before the step change.

“…For example, ammonium has been shown to inhibit the growth of mouse L cells' and canine kidney (MDCK) cells. 4 Ammonium also inhibits the production of a number of viruses'-'' as well as interferon.". '* Lactate accumulation can lower the pH of the culture, subsequently inhibiting growth, since mammalian cells grow optimally over narrow pH ranges.''…”

Section: Introductionmentioning

confidence: 99%

Mathematical descriptions of hybridoma culture kinetics: I. Initial metabolic rates

Glacken

Adema

Sinskey

1988

182

The lack of quantitative descriptions of mammalian culture kinetics limits the ability to optimally design and control cell culture bioreactors. This limitation is ad dressed by developing mathematical equations relating the initial growth rate and the antibody productivity of the hybridoma cell line, CRL-1606, to its environmental state. This initial rate approach, in contrast with steady state analysis of chemostat cultures, approximates steady state behavior, since the rates were measured over brief time intervals and at low cell concentrations (< 50,000 cells/mL). The advantage of this approach is that it is much faster than the chemostat approach.An equation for the growth rate was developed that superimposed Monod equations in serum and glutamine with "noncompetitive" -type inhibition constants were inversely proportional to the lactate and ammonium concentrations. The Monod constant is critical for relating initial, low cell level culture states to other states.Lactate was found to be the only environmental parameter that significantly inhibited antifibronectin monoclonal antibody production by the CRL-1606 hybridomas. Volumetric productivity was strictly related to culture viability, which was observed to decline at growth rates below 0.02 h(-1). Lactate was also found to significantly inhibit ammonium production.