Figure 4

Test of optimized antibody expression vectors in HEK293-6E cells. Three different vectors pCMV-, pCMV-oriP1- and pCMV-neo^- -scFv-Fc-4E3 encoding a CD30 specific scFv-Fc antibody, and another three vectors pCMV-, pCMV-oriP1- and pCMV-neo^- -scFv-Fc-RNase-4E3 encoding a CD30 specific immunoRNase, respectively, were transiently transfected into HEK293-6E cells in 50 mL scale in shake flasks. Production of each vector and each construct were performed as triplicates. A) After 48 and 72 h, volumetric yields of scFv-Fc and scFv-Fc-RNase were tested from production supernatants by human IgG/Fc capture ELISA. B-C) After 72 h of scFv-Fc (D, triplicate samples individually purified) and scFv-Fc-RNase (E, triplicate samples combined prior purification) were purified by protein A affinity chromatography and volumetric yields were calculated. Percentage recovery is calculated in comparison to the volumetric yields measured from supernatants (A). D) Several production parameters were tested like transfection efficiency (co-transfection with an GFP expressing vector), viable cell number and viability as well as E) concentrations of glucose, L-lactate, L-glutamine and L-glutamate.