Current Choices of Host Cells in Biotech Bacteria Cells Yeast Transgenic Animals Transgenic Plants Animal Cells
Comparison of Monoclonal Antibody Produced from CHO & Transgenic Goats Assumption Annual Yield (Kg/yr) Batch Yield (grams/L) 60 goat herd 350 L/animal year 40 5.0 Transgenic Goats Grange Castle 6 X 12,500 L Bioreactors 4000 3.4 CHO Bioreactor
The Majority of Biotech Products on the Market Are Made in Animal Cells
Comparison of Animal and Microbial Culture 10 6 cells/mL 10 9 -10 10 cells/mL Growth density 10 5 cells/mL 1 cell Seeding density 10000-100000 nm 100-2000 nm Size Very susceptible Less affected Environmental FX Key for buffering Sometimes CO 2 Requirement Complex Usually simple Nutritional Rqmt Low High O 2 Requirement 1-5% per hour 10-50% per hour Growth Rate Present Present Cell membrane Generally absent Generally present Cell wall Animal Cells Microbes Features
A closed culture system which contains an initial, limited amount of nutrient. The inoculated culture will pass through a number of phases following a growth curve. The growth curve contains four distinct regions as
Cells have adjusted to their new environment The cells are dividing at a constant rate resulting in an exponential increase in the number of cells present. This is known as the specific growth rate and is represented mathematically by first order growth rate
dX = (μ – kd) X
where X is the cell concentration,
μ is the cell growth rate
kd is the cell death rate.
The cell death rate is sometimes neglected if it is considerably smaller than the cell growth rate.
Monod batch kinetics is represented mathematically in the following equation:
μ = μ max S
where μ is the specific growth rate, μ max is the maximum specific growth rate, S is the growth limiting substrate concentration and Ks is the saturation constant which is equal to the substrate concentration that produces a specific growth rate equal to half the max specific growth rate
For Primary Metabolite production conditions to extend the exponential phase accompanied by product excretion
For Secondary Metabolite production, conditions giving a short exponential phase and an extended production phase, or conditions giving a decreased growth rate in the log phase resulting in earlier secondary metabolitwe formation.
While cells are still growing exponentially, but nutrients are becoming depleted, concentrated feed medium (usually a 10-15 times concentrated basal medium) is added either continuously (as shown) or intermittently to supply additional nutrients, allowing for a further increase in cell concentration and the length of the production phase.
In contrast to an intermittent-harvest strategy, fresh medium is added proportionally to cell concentration without any removal of culture broth.
To accommodate the addition of medium, a fedbatch culture is started in a volume much lower than the full capacity of the bioreactor
In batch cultures and most fedbatch processes, lactate, ammonium, and other metabolites eventually accumulate in the culture broth over time, affecting cell growth, glycoform of the product and productivity.
Other factors, such as high osmolarity and accumulation of reactive oxygen species, are also growth inhibitory
After extended exposure to low glucose concentrations, cell metabolism is directed to a more efficient state, characterized by a dramatic reduction in the amount of lactate produced. Such a change in cell metabolism from the normally observed high lactate producing state to a much reduced lactate production state is often referred to as metabolic shift.
Very high cell concentrations and product titers were achieved in hybridoma cells.