Figure 5

Prolonged transient protein production in HEK293-6E cells. A) and B) The vectors pCMV-oriP1, pCMV-neo^- and pCMV containing CD30 specific scFv-Fc- and scFv-Fc-RNase-4E3 constructs, respectively, were transiently transfected into HEK293-6E cells in 25 mL scale in shake flasks. In addition, pCMV2.2-scFv-Fc with the MUC1-specific affinity maturated scFv HT186-D11 [8] and a plasmid pCS2-venus expressing the yellow fluorescent reporter protein (YFP, venus) were transfected in parallel attempts. Production samples were taken 3, 6, 7, and 8 days after transfection. A) Volumetric yields of scFv-Fc and scFv-Fc-RNase were tested by gel densitometry. C) SDS-PAGE was performed with 5 μL of each supernatant under reducing conditions. Gels were stained with coomassie (G250). Purified scFv-Fc- and scFv-Fc-RNase-4E3 were used as standards for gel densitometry. YFP was compared to the scFv-Fc standard.