FIG. 4.
Release of HA and M1 proteins expressed by vTF7-3 infection/transfection in CV1 and HeLa T4 cells. HA and M1 were expressed in CV1 (A) or HeLa T4 (B, upper) cells by vTF7-3-expressed, T7-RNA polymerase-driven expression, according to the protocol of Gómez-Puertas et al. (18). At 60 h p.t., the culture medium was harvested and pelleted through a 30% sucrose cushion and cells were prepared as described in Materials and Methods. Where indicated, exogenous bacterial NA (exo NA) was added to the replacement medium at 20 h p.t. HeLa T4 cells (B, lower) were also transfected with HA, NA, and M1 protein expression plasmids, and the culture media and cells were analyzed as described above. Samples were analyzed by SDS-PAGE followed by immunoblotting to detect viral proteins.