FIG. 2.

Production of adenoviral vectors pseudotyped with transiently expressed fiber proteins by using the transient transfection/infection system. (A) The fiber-deleted adenoviral vector Ad5.βgal.ΔF can be grown in packaging cell lines transiently or stably expressing different fiber proteins to generate Ad5.βgal.ΔF/F⁺ adenoviral particles containing fiber. The adenoviral vector is used to infect 293T cells that have been transfected with a fiber expression plasmid. The resulting particles will have new receptor tropisms dependent on the fiber protein expressed or on the peptide ligand that was inserted into the HI loop of the fiber protein. (B) Western immunoblot analysis of the Ad5 fiber and penton capsid proteins. An equivalent number of adenovirus particles for the indicated vectors were subjected to denaturing SDS-PAGE and Western immunoblot analysis. For detection of the fiber protein incorporated onto the adenoviral capsid, the membrane was developed with the rabbit anti-fiber polyclonal antibody and an anti-rabbit IgG horseradish peroxidase-conjugated secondary antibody by chemiluminescence. For detection of the penton capsid protein, the same membrane was developed with the rabbit anti-penton polyclonal antibody and an anti-rabbit IgG horseradish peroxidase-conjugated secondary antibody by chemiluminescence.