Figure 5.
Figure 5. The effect of disruption of the 14-3-3ζ binding sites in GPIbα and GPIbβ by mutagenesis on the VWF binding function. (A) CHO cells expressing wild-type GPIb-IX (1b9), S609A, S166A, or S166A/S609A mutants were incubated with human VWF in the presence of ristocetin or with ristocetin alone (controls). VWF binding to cells was analyzed by flow cytometry using FITC-labeled anti-VWF antibody, SZ29. The same cells were also analyzed for surface expression of GPIb-IX by flow cytometry using an anti-GPIbα antibody SZ2 followed by a FITC-labeled goat anti-mouse IgG. Results from a typical experiment are shown in panel A. Quantitative data are shown in panel B, in which fluorescence intensity of VWF binding (mean ± SD, 6 experiments) was corrected for the relative level of GPIb-IX expression. FL indicates fluorescence.

The effect of disruption of the 14-3-3ζ binding sites in GPIbα and GPIbβ by mutagenesis on the VWF binding function. (A) CHO cells expressing wild-type GPIb-IX (1b9), S609A, S166A, or S166A/S609A mutants were incubated with human VWF in the presence of ristocetin or with ristocetin alone (controls). VWF binding to cells was analyzed by flow cytometry using FITC-labeled anti-VWF antibody, SZ29. The same cells were also analyzed for surface expression of GPIb-IX by flow cytometry using an anti-GPIbα antibody SZ2 followed by a FITC-labeled goat anti-mouse IgG. Results from a typical experiment are shown in panel A. Quantitative data are shown in panel B, in which fluorescence intensity of VWF binding (mean ± SD, 6 experiments) was corrected for the relative level of GPIb-IX expression. FL indicates fluorescence.

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