Fig. 2.

Expression of spike protein in infected cells and NDV particles. (a) Expression of the S and S-F protein in infected cells. Vero E6 cells were infected with three NDV vectors encoding the S or S-F for 16 to 18 h. A WT NDV control was included. The next day, cells were fixed with methanol-free paraformaldehyde. Surface proteins were stained with anti-NDV rabbit serum or a spike receptor-binding domain (RBD)-specific monoclonal antibody CR3022. A scale bar of 200 µM were shown. (b) Incorporation of S and S-F into NDV particles. Three NDV vectors expressing the S or S-F including the NDV_LS_S (green), NDV_LS_S-F (red) and NDV_LS/L289A_S-F (blue) were concentrated through a 20% sucrose cushion. Two clones were shown for NDV_LS_S and NDV_LS_S-F. The concentrated WT NDV expressing no transgenes was used as a control. Two micrograms of each concentrated virus were resolved on a 4–20% SDS-PAGE, the spike protein and NDV HN protein were detected by western blot using an anti-spike 2B3E5 mouse monoclonal antibody and an anti-HN 8H2 mouse monoclonal antibody.