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. 2023 Jul 22;15(14):3721. doi: 10.3390/cancers15143721

Figure 3.

Figure 3

Binding affinities of the engineered chimeric and humanized 2EF. (A) ELISA binding assay of Ch2EF (batches A, B), Hu2EF-4, Hu2EF-6, Hu2EF-7 to recombinant Trop-2-Fc chimera immobilized to the substrate. Murine 2EF was used as a benchmark. Trop-2-Fc was used at 1 µg/mL (left) or at 0.1 µg/mL (right). Absorbance values (Y-axis) are plotted at each tested Ab concentration (X-axis). (B) Data from competitive flow cytometry analysis of murine 2EF and Ch2EF binding to MTE4-14/Trop-2 transfectants were plotted as the ratio of unlabeled mAb/labeled mAb versus mean fluorescence intensity (left) and as labeled mAb/total mAb ratio versus mean fluorescence intensity (right). Theoretical binding curves are in black. (C) Data from competitive flow cytometry analysis of Colo205 colon cancer cells stained with murine 2EF-Alexa488, as competed-out by unlabeled engineered 2EF variants: (left) Ch2EF, Hu2EF-5, Hu2EF-6 (batches 1 and 2); (right) Hu2EF-7 produced in CHO-K1 or YB2/0 low-fucosylation cells.