| Popis: |
Growth and monoclonal antibody production kinetics of the hybridoma cell line NB1 were studied in batch, chemostat and two-stage chemostat culture. Growth associated production kinetics were observed in batch culture when glutamine was the first nutrient to be depleted. The growth phase was extended and production increased by supplementing the culture with extra glutamine, essential and non-essential amino acids. Monoclonal antibody was produced by the viable cell population with no evidence for its release from lysing cells. Increasing the concentration of individual amino acids in batch culture appeared to have a negative effect on cell growth and in some cases a positive effect on antibody production. The effect on production may have been indirect, due to the effect on growth rate. Under conditions of leucine and isoleucine limitation the kinetics of antibody production in batch culture were changed to growth dissociated with the majority of the antibody being produced during the decline phase of the culture. The kinetics of production were therefore dependent on the nutrient limiting growth. In single-stage chemostat culture monoclonal antibody production was negatively growth rate associated, in contrast to the positively growth associated production generally observed in batch culture. Therefore the feasibility of optimising negatively growth associated antibody production in the second stage of a two-stage chemostat was studied. Steady states in terms of cell concentration, nutrient utilisation and antibody production were observed in both stages of a two-stage chemostat. Monoclonal antibody production in both stages of a two-stage chemostat was negatively growth rate associated. Specific production rate increased with decreasing dilution rate. The specific production rate in the second stage was less than the first stage at an equivalent dilution rate even though the growth rate was less, except at dilution rates below 0.02 h-1 when the growth rate in stage two was negative. Feeding the second stage of a two-stage chemostat with amino acids increased the growth rate but decreased the specific production rate. Attempts to decrease the growth rate of a fed second stage by increasing the volume and therefore decreasing the dilution rate relative to the first stage were unsuccessful. Growth rate remained high, however volumetric production was significantly increased. The concentration of an amino acid in the medium appeared to influence its rate of utilisation in batch and chemostat culture. Increasing the concentration of an individual amino acid tended to have a negative effect on cell growth. Decreasing the concentration of an individual amino acid reduced the extent to which it was utilised. Increasing the concentration of all amino acids increased their rate of utilisation without increasing the cell concentration. |
