NADPH concentration in cell culture fluids and cell lysate
The CHO cell cultures from five different recombinant molecules were centrifuged at 800 x g for 10min at room temperature. The supernatants were sampled and removed. The cell pellets were re-suspended to their original volumes in 50 mM HAc-NaAc buffer and followed by 10000 x g for 30min at 4oC. In total, the five supernatants of the CFFs and the five cell lysates were obtained. Their NADPH concentrations were measured and the results were depicted in Figure 5. Among the five supernatants of the CFFs, the highest NADPH concentrations was 0.1 µM and the two of them were below the quantification limit (0.01µM). The observation suggested that the NADPH concentrations were low in the CFFs. The NADPH concentrations in the five cell lysates varied from 0.1 µM to 2.4 µM. The variability of NADPH concentration was large both in the CFFs and in the cell lysates. The NADPH concentration in the cell lysates was much higher than that in the CFFs. This partially explained why the bisulfite bond reductions only occurred in some manufacturing processes and why the harvest process could impact their occurrence for a given CCF.