Figure 5

Binding characteristics of PH1 derivatives. A, B, ELISA on coated mGroEL-MUC1 peptide. Increasing concentrations of PH1 Fab (circles), bibody (squares) and tribody (triangles) purified from NS0 (A) or P. pastoris cell cultures (B) were allowed to interact with coated mGroEL-MUC1. After 405 nm OD readout, saturation binding curves and constants were calculated using non-linear regression. Shown is the mean ± SD of triplicates. C, Binding of PH1 derivatives to cell associated MUC1. Histograms of flow cytometric analysis showing the interaction with MUC1⁺ OVCAR3 cells of the PH1 bibody and tribody from NS0 or P. pastoris origin. Cells were stained with 20 μg/ml PH1 bibody or tribody, followed by mouse anti-human κ (filled) or anti-c-myc IgG₁ (open) isotype control MoAb. Bound MoAbs were detected with Alexa 488-conjugated anti-mouse IgG.