Using cell size kinetics to determine optimal harvest time for Spodoptera frugiperda and Trichoplusia ni BTI-TN-5B1-4 cells infected with a baculovirus expression vector system expressing enhanced green fluorescent protein

Infecting insect cells with a baculovirus expression vector system (BEVS) is an increasingly popular method for the production of recombinant proteins. Due to the lytic nature of the system, however, determining the optimal harvest time is critical for maximizing protein yield. We found that measuring the change in average diameter during the progress of infection with an automated cell analysis system (Cedex HiRes, Innovatis AG) could be used to determine the time of maximum protein production and, thus, optimal harvest time. As a model system, we use insect cells infected with a baculovirus expressing enhanced green fluorescent protein (EGFP). We infected two commonly used insect cell lines, Spodoptera frugiperda (Sf-9) and Trichoplusia ni BTI-TN-5B1-4 (Hi5) with an Autographa californica nuclear polyhedrosis virus (AcNPV) encoding EGFP at various multiplicities of infection (MOI). We monitored the progress of infection with regard to viability, viable cell density and change in average cell diameter with a Cedex HiRes analyzer and compared the results to the EGFP produced. Peak protein production was reached one to two days after the point of maximum average diameter in all conditions. Thus, optimal harvest time could be determined by monitoring the change in average cell diameter during the course of an infection of a cell culture

On-line monitoring of infected Sf-9 insect cell cultures by scanning permittivity measurements and comparison with off-line biovolume measurements

Two infected Sf-9 cell cultures were monitored on-line by multi-frequency permittivity measurements using the Fogale BIOMASS SYSTEM® and by applying different off-line methods (CASY®1, Vi-CELL™, packed cell volume) to measure the biovolume and the mean diameter of the cell population. During the growth phase and the early infection phase the measured permittivity at the working frequency correlated well with the different off-line methods for the biovolume. We found a value of 0.67 pF cm−1 permittivity per unit of total biovolume (CASY) (μL mL−1). After the maximum value in the permittivity was reached, i.e. when the viability of the cultures decreased significantly, we observed different time courses for the biovolume depending on the applied method. The differences were compared and could be explained by the underlying measurement principles. Furthermore, the characteristic frequency (fC) was calculated from the on-line scanning permittivity measurements. The fC may provide an indication of changes in cell diameter and membrane properties especially after infection and could also be an indicator for the onset of the virus production phase. The changes in fC were qualitatively explained by the underlying equation that is correlating fC and the properties of the cell population (cell diameter, intracellular conductivity and capacitance per membrane area)

Bioreactor technology for sustainable production of plant cell-derived products

The successful cultivation of plant cell and tissue cultures for the production of valuable chemical components requires the selection of an appropriate bioreactor. Selection criteria are determined based on a number of factors that are intrinsic to particular plant cell or tissue cultures and are influenced by the process objectives. Due to the specific properties of plant cell and tissue cultures, bioreactor systems may differ significantly from those used for microorganism or animal cell cultures. Furthermore, the differences from one plant culture to another can be immense; it is obvious that the optimal bioreactor system for a plant suspension cell culture is different to one for a plant tissue culture in many ways. General considerations are presented, and based on these key points, selection criteria are used to establish a “bioreactor chooser” tool. The particular details of the most relevant bioreactor types for plant cell and tissue cultures are listed and described. To produce valuable products, the process also needs to be scaled up to an economically justifiable size, which is usually done either by scaling up the size of the bioreactor itself or by bioreactor parallelization. Therefore, the most significant influencing factors are also discussed