Subpassaging cells on microcarriers: the importance for scaling up to production.
Optimization of microcarrier processes is dependent upon efficient, serial subcultivation routines. Established methods have been modified for a high degree of cell detachment from microcarriers, and transfer of a maximum number of viable cells from one culture to the next during the scale up process. Cultures of MRC-5 and Vero cells were studied, and cell inocula were obtained from different growth phases (i.e. exponential versus stationary) to investigate growth in subsequent cultures. Microcarrier cultures containing confluent cells were washed with EDTA-PBS and then exposed to trypsin (185 U/ml) in PBS (pH 8.0, 37 degrees C). 95-100% of the cells detached from the microcarriers with a viability greater than 95% following a 10 minute exposure to the trypsin. The presence of residual trypsin in the inoculum was investigated with respect to subsequent growth, and no significant effect was found. The methods developed at the laboratory scale (0.25 to 1.5 l cultures) were successfully applied to pilot scale (1 to 100 l cultures), and resulted in split ratios of up to 1:10 for MRC-5 cells and 1:100 for Vero cells. These results show that the modified subpassaging method and an optimal cell inoculum are vital in establishing efficient, industrial scale microcarrier processes.
D000818:Animals; D001709:Biotechnology / Q000379:methods; D002448:Cell Adhesion; D002455:Cell Division; D002470:Cell Survival; D002478:Cells, Cultured / Q000502:physiology*; D003911:Dextrans; D006801:Humans; D008863:Microspheres; D014357:Trypsin; D014709:Vero Cells
D003911:Dextrans; D014357:Trypsin