Generating Antibodies to the Gene 3 Proteins of Infectious Bronchitis Virus

Pendleton, Amanda R.; Machamer, Carolyn E.

Infectious bronchitis virus (IBV), a group 3 coronavirus, produces three proteins (IBV E, IBV 3a, and IBV 3b) from subgenomic mRNA 3 during infection. IBV E, a viral envelope protein, plays a role in virus budding, possibly by altering membrane morphology at the virus assembly site. In addition to this role, IBV E may also function as a viroporin, although no data from infected cells have confirmed this possibility definitively. Conversely, the IBV 3a and IBV 3b proteins are nonstructural proteins. These proteins are dispensable for replication in cell culture, but are thought to be important for infection of the natural host. This chapter details methods for generating and screening antibodies to these gene 3 proteins. Antibodies were raised in rabbits following inoculation with IBV-specific peptides and GST fusion proteins, and were screened by immunofluorescence, radioimmunoprecipitation, and immunoblotting.