Analysis of interferon gamma protein expression in zebrafish (Danio rerio)

• A monoclonal antibody to zebrafish IFN-γ has been produced and characterised.
• The moAb detected induced IFN-γ protein expression following PHA stimulation of kidney/spleen leukocytes.
• Antigen specific induction of IFN-γ protein in zebrafish following immunization and re-stimulation was also demonstrated.
• A positive correlation between mRNA and protein levels of IFN-γ in zebrafish was revealed in this study.

IFN-γ is a major effector cytokine, produced to induce type I immune responses. It has been cloned in several fish species including zebrafish, however to date few studies have looked at IFN-γ protein expression and bioactivity in fish. Hence, the current study focused on developing a monoclonal antibody (moAb) against zfIFN-γ. We show that the zfIFN-γ moAb specifically recognises E. coli produced recombinant IFN-γ protein and zfIFN-γ produced in transfected HEK293 cells, by Western blot analysis. Next we analysed the production of the native protein following expression induced by PHA stimulation of leukocytes in vitro or antigen re-stimulation in vivo. We show the IFN-γ protein is produced as a dimer, and that a good correlation exists between transcript expression levels and protein levels.