Modified monoclonal antibody production kinetics kappa/gamma mRNA levels, and metabolic activities in a murine hybridoma selected by continuous Culture.

During long-term continuous culture of the hybridoma cell line 11317, a better-producing subclone (I1317-SF11), giving improved productivity, has been selected. The comparison of the original cell line (I1317-DC) with this subclone revealed that although the growth patterns of both clones were similar, both in continuous and in batch cultures, considerable differences could be seen between the clones with respect to monoclonal antibody (MAB) accumulation, MAB production rate, the levels of mRNA coding for heavy and light chains of IgG, and some metabolic activities. In continuous culture as well as in batch culture, I1317-SF11 showed increased levels of mRNA coding for kappa and gamma chains compared with I1317-DC and/or a modified ratio of the mRNA species when compared to that in I1317-DC. Using pulse experiments, it could be established that the biosynthesis of both chains was augmented in I1317-SF11. Although the kappa and gamma mRNA levels were modified or inversed for I1317-SF11, the cells always synthesized more kappa than gamma chains. The overall increase in the synthetic activity of I1317-SF11 is suggested as one reason for the considerable increase of IgG productivity and product accumulation in continuous culture as well as in repeated batch cultures. Tests concerning metabolic activity revealed that I1317-SF11 had a predominantly glycolytic metabolism independent of growth requirements, whereas for I1317-DC the metabolism became increasingly glycolytic with increased growth. The antibody yield coefficient of I1317-SF11 on glutamine was significantly higher than that of I1317-DC for the continuous culture, whereas the antibody coefficients on glucose were almost similar for both clones under the different culture conditions used. Both antibody coefficients were considerablly influenced by the specific growth rate.All these facts together lead to the conclusion that subclone I1317-SF11 uses more of the energy available, or it was the energy and/or precursors available for the synthesis and production of MAB more efficiently than the thesis and production of MAB more efficiently than the original cell line. Although the levels of mRNA coding for heavy and light chains of IgG were modified, it could be confirmed that the overall regulation of MAB-synthesis and -production occurs post-translationally and that at higher growth rates, more biosynthetic activity is diverted to biomass production.

A new method for the encapsulation of mammalian cells.

A new encapsulation method was developed for the cultivation of mammalian cells. The capsules were produced using a solution of sodium cellulose sulphate (CS)(1.5%) and poly-dimethyl-diallyl-ammonium chloride (PDMDAAC). When CS droplets fell into the precipitation bath consisting of a 2% solution of PDMDAAC, immediately a membrane at the interphase was built up. The influences of varying encapsulation process parameters on capsule characteristics, cell growth, and monoclonal antibody production were tested. This new method showed advantages when compared to other methods mainly due to time simplicity of the whole process.

Application of sensors for the determination of physical and biochemical parameters in the fermentation of animal cells.

Biosensors, sondes, and transducer elements are reviewed with respect to the application for fermentation processes of animal cells. Hereby, the possible connection of these elements to the fermentor is shown. Specially, the (possible) online determinations of the viable and total cell count, of the glucose, the lactate, the pyruvate, and the ammonia concentration in the medium of fermentors are presented. Finally, some sensors for the on-line estimation of the product concentration are described.

Use of a dynamic filtration method for separation of animal cells.

For solving the problem of concentrating and washing of viable cell suspensions a dynamic filter device has been evaluated. Results show that concentrating of cells under sterile conditions can be performed. With a laboratory scale filter (Sulzer-Biodruckfilter BDF-01) a 9-fold concentration was achieved, the filtration speed was about 15 l/h. The viability did not change significantly. Details of experiments are given and advantages are discussed.

Invasive infrared sensor for the determination of the cell number in a continuous fermentation of hybridomas.

A new and simple device for the on-line determination of the total cell count in fermentation vessels is presented. It is based on fibre optic techniques, produces a constant infrared signal and determines the cell count via the scattered light. The linear range between the cell count and the infrared response lies between 1 X 10(5) and more than 2 X 10(6) cells per ml. This sensor shows good correlation between the cell count and the infrared response in fermentations with cell viability of more than 80%. The feeding pumps in these fermentations can be controlled via this sensor to get fermentation with stationary cell counts.