Figure 3.

(a) C¹⁴ autoradiogram of SDS-PAGE gel showing Trastuzumab IgG expression in XpressCF+^® with either co-expression of the HC and LC or expression of the HC alone in the presence of the PFLC reagent protein. The HC/LC co-expression shows a plasmid ratio titration, which is required to identify the ratio of the two proteins which yields optimal IgG titer and assembly. The concentration of PFLC reagent was also titrated in the HC alone with PFLC expressions. (b) Trastuzumab IgG titer calculated from the autoradiogram in part (a). Green bars represent the HC/LC co-expression while blue bars represent the HC expression alone in the presence of the PFLC reagent. Error bars are calculated from SD of acid precipitable C14 detected by liquid scintillation counter (n = 2) (c) Comparison of the relative improvement of cell-free expression by C¹⁴ observed with the PFLC reagent compared to HC/LC co-expression for four mAbs and one bispecific antibody (FabFc:scFvFc format). The gray bars represent the theoretical titer improvement expected given a constant level of cell-free protein synthesis, folding, and assembly between the two reaction formats. Orange bars represent the additional titer increase observed, which is partially attributed the LC acting as a co-chaperone to improve HC solubility and final IgG folding and assembly.