Problems with serum-free production of antithrombin III regarding proteolytic activity and product quality

Serum- and protein-free media formulations for the Chinese hamster ovary cell line DUKXB11

“…The detection limit of our L-LDH assay is ≈0.0025 units/ml in conditioned medium, and from this we estimate that during a 24 h conditioning phase less than 1% of the cells, if any, lysed. These results are in sharp contrast to those reported by Teige et al (1994). Teige et al (1994) used CHO cells that were not weaned off serum for their serum-free culture, whereas in our study a careful adaptation of our CHO cell lines to serum-free conditions and a medium optimization took place.…”

“…These results are in sharp contrast to those reported by Teige et al (1994). Teige et al (1994) used CHO cells that were not weaned off serum for their serum-free culture, whereas in our study a careful adaptation of our CHO cell lines to serum-free conditions and a medium optimization took place. These different approaches to the serumfree cultivation process can explain the dramatic differences in the sensitivity of CHO cells to serum-free conditions observed in both studies.…”

“…In contrast, in medium UC203, in which CHO cells also had a generation time of ≈12 h, a lag-phase of about 48 h was observed (Ii et al, 1995). Teige et al (1994) reported that cell lysis can be a major problem when ATIII is produced in serum-free media. This prompted us to investigate the degree of cell lysis in medium CHO-T1-SF after inoculation at higher cell densities (2·10 4 cells/cm 2 ).…”

Mammalian cell-produced therapeutic proteins: heterogeneity derived from protein degradation

“…In the case of EPO, high glutamine concentration (16-20 mM) in the culture medium inhibited the protease activity which suggested that amino acid starvation (Glu, Asp, Asn and Ser) induced protease activity [55,56]. Protease activity may be minimized in cell cultures supplemented with fetal bovine serum which contains significant amount of protease inhibitors [57]. However, regulatory requirements necessitate the use of animal protein-free media formulations for production of recombinant therapeutic proteins.…”

Fermentor temperature as a tool for control of high-density perfusion cultures of mammalian cells

“…One possible effect is the decrease of the proteolytic activity in the fermentor (Enfors, 1992;Kratje et al, 1994;Teige et al, 1994). This might be essential if complex proteins prone to degradation are produced by cell lines with high proteolytic activity, such as BHK (Adamson, 1994;Kratje et al, 1994;Teige et al, 1994).…”