Spectrophotometry method for titration of trout interferon, and its application to rainbow trout fry experimentally infected with viral haemorrhagic septicaemia virus

Abstract

The activity of a reference preparation of rainbow trout circulating interferon (IFN), induced by experimental infection of fish with viral haemorrhagic septicaemia virus, was detected on the basis of its non-specific protective effect for RTG-2 cells, grown in microplates and challenged with infectious pancreatic necrosis virus. The IFN was titrated by spectrophotometric assessment of the absorbancy (wavelength 595 nm) of dried cell monolayers, stained with crystal violet after virus challenge. By definition, the reciprocal of the IFN sample dilution giving a cell layer with 50% of the dye absorbancy of the uninfected control cell layer represented the IFN titre. This spectrophotometric method of determining IFN titre appeared as sensitive as the plaque assay normally used for this purpose, but was better suited to the processing of large numbers of IFN samples, required by investigations on the pathogenesis of fish viroses. The method proved effective at detecting IFN in homogenates of whole rainbow trout fry, thus allowing individual screening for IFN of small fish which are the usual targets of systemic viroses.