Effect of Peptones and Study of Feeding Strategies in a CHO Based Fed-batch Process for the Production of a Human Monoclonal Antibody

Chotteau, Véronique; Wahlgren, C.; Pettersson, Helena

doi:10.1007/978-1-4020-5476-1_61

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“…For many years, there has been an increasing demand in developing efficient production processes for biopharmaceuticals like monoclonal antibodies (Chotteau et al, 2007;Kunert et al, 2000), recombinant proteins (Baker et al, 2001;Gaillet et al, 2007), and viral vaccines (Genzel et al, 2004;Lohr et al, 2009) using animal and human cell lines as cell substrate. However, a major problem that can be seen with almost all the producer cell lines is the highly inefficient use of nutrients.…”

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confidence: 99%

Measurement of key metabolic enzyme activities in mammalian cells using rapid and sensitive microplate‐based assays

Janke

Genzel

Wahl

et al. 2010

Biotech & Bioengineering

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Sensitive microplate-based assays to determine low levels of key enzyme activities in mammalian cells are presented. The enzyme platform consists of four cycling assays to measure the activity of 28 enzymes involved in central carbon and glutamine metabolism. The sensitivity limit of all cycling assays was between 0.025 and 0.4 nmol product. For the detection of glutaminase activity, a new glutamate cycle system involving the enzymes glutamate dehydrogenase and aspartate transaminase was established. The relative standard deviation of the method was found to be 1.7% with a limit of detection of 8.2 pmol and a limit of quantitation of 24.8 pmol. Hence, cell extracts could be highly diluted to reduce interferences caused by other components in the extract, which in addition minimized underestimates or overestimates of actual enzyme activities. Since substrate concentrations could be maintained at a nearly constant level throughout the assay product accumulation during the reaction was low, which minimized product inhibition. As an example, the enzyme platform was used to investigate maximum enzyme activities of stationary-phase MDCK cells grown in serum-containing GMEM medium as typically used in influenza vaccine production.

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