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. 2018 Jan 10;9(17):13206–13221. doi: 10.18632/oncotarget.24152

Figure 4. ADCC activity of afucosylated ch2448.

Figure 4

An afucosylated variant of ch2448 (aF-ch2448) was generated. (A) Antibodies ch2448 and aF-ch2448 (and human IgG control) were run on SDS-PAGE in non-reducing conditions. Coomassie Blue staining of the gel showed antibodies at ∼150 kDa in size. Western blotting of samples run in-parallel showed that the Aleuria Aurantia Lectin was not able to recognize aF-ch2448, demonstrating the loss of core fucose. (B) The binding of aF-ch2448 (Mutant) retained similar specificity of ch2448 on live IGROV1 cells by flow cytometry. Cells were incubated with ch2448 or af-ch2448 at various concentrations. Binding was measured by an increase of the normalized mean fluorescence intensity (nMFI). Results were representative of three independent experiments. (C) ADCC activity of ch2448 and aF-ch2448 was measured on OVCAR3, IGROV1, SKOV3, MC7-D10 and MCF7-2101 cell lines. ADCC activity was measured as fold induction of the NFAT pathway. Values are means ± standard deviations of triplicates. Similar results were obtained in 3 independent experiments.