4.1 Enzymatic factors.
Researchers found that during the protein harvest operation, high mechanical shear force could cause cell lysis. Consequently, more intracellular enzymes are released to the Harvest Cell Culture Fluid (HCCF) thus lead to a higher level of disulfide bond reduction (Hutterer et al., 2013; Trexler-Schmidt et al., 2010). Alternatively, low dissolved oxygen (DO) levels in cell culture or HCCF can also cause disulfide bond reduction since there is a limited amount of oxygen molecules to consume NADPH, thus leaving enough NADPH as “electron fuel” for the disulfide bond reduction reaction (Mun et al., 2015; O’Mara et al., 2019). These aspects of root causes have been the focus of recent industrial studies.