Fig. 3.
Cell-based binding assay for the interaction of S protein with ACE2. BHK-SΔ18 cells were grown to monolayers in the presence of the fluorescent fatty acid BODIPY FL C12. After detachment by treatment with Accutase, cells were used for overlay assays. (A) Confluent Vero E6 cells were overlaid with PBS (a and b), BHK-SΔ18 (c and d), BHK-SΔ18 that had been pretreated with anti-SARS serum (e and f), or with fluorescence-labeled BHK21 cells (g and h). Phase-contrast pictures are shown in panels on the left side (a, c, e, g); pictures obtained by fluorescence microscopy are shown on the right (b, d, f, h). (B) Confluent Vero E6 cells were treated with different concentrations of mβCD. In cholesterol replenishment experiments, cells were incubated in the presence of cholesterol. After these treatments, fluorescence-labeled BHK-SΔ18 cells were added. Bound cells were detached by trypsin treatment and subjected to flow cytometry. The figures show a representative image. The bars represent the mean ± standard deviation of 4 independent experiments.