Cloning, expression and immunoassay detection of ferret IFN-γ

Abstract

Ferrets (Mustela putorius furo) develop symptoms upon influenza infection that resemble those of humans, including sneezing, body temperature variation and weight loss. Highly pathogenic strains of influenza A, such as H5N1, have the capacity to cause severe illness or death in ferrets. The use of ferrets as a model of influenza infection is currently limited by a lack of species-specific immunological reagents. Interferon gamma (IFN-gamma) plays a key role in the development of innate and adaptive immunity and the regulation of Th1-type immune responses. Here we describe the cloning of the full-length cDNA for ferret IFN-gamma. Multiple sequence alignment of the predicted amino acid sequence with those of other species indicates that the predicted ferret protein shares the highest identity with Eurasian badger IFN-gamma. We raised two hybridoma clones expressing monoclonal antibodies against recombinant ferret IFN-gamma capable of detecting IFN-gamma protein derived from mitogen-stimulated ferret PBMCs by immunoblotting, ELISA and ELISPOT assay. Finally, an ELISA utilizing the ferret-specific antibodies detected elevated levels of IFN-gamma in serum samples from H3N2 influenza A-infected ferrets.

A Single Immunization with CoVaccine HT-Adjuvanted H5N1 Influenza Virus Vaccine Induces Protective Cellular and Humoral Immune Responses in Ferrets

“…Recently, monoclonal antibodies specific for human cluster-of-differentiation (CD) antigens that cross-react with those of ferret lymphocytes have been defined (35,43). Antibodies specific for ferret gamma interferon (IFN-␥) that can be used to detect the production of this cytokine by ferret T cells (for example, by enzyme-linked immunospot [ELISPOT] assay) also have been produced (28). However, although reagents are becoming available, assays for the detection of virus-specific T cells have not been described so far to our knowledge.…”

Contemporary Seasonal Influenza A (H1N1) Virus Infection Primes for a More Robust Response To Split Inactivated Pandemic Influenza A (H1N1) Virus Vaccination in Ferrets

“…A number of recent publications have, however, described the development of ferret-specific reagents as well as a number of cross-reactive reagents used in other systems (17,19). In this study, we utilized an anti-human CD8 antibody to assess the role of memory CD8 ϩ T cells in withinsubtype heterologous protection.…”

“…Table 1 VOL. 17,2010 PRIMING OF THE pMIV RESPONSE BY SEASONAL H1N1 EXPOSURE 1999 received two doses of pMIV 3 weeks apart. The fourth group (T/M) received a single dose of TIV (T) and then a single dose of pMIV.…”

The current state of H5N1 vaccines and the use of the ferret model for influenza therapeutic and prophylactic development

“…As well, the characterization of ferret cytokines and immune molecules, essential components to the evaluation of therapeutics and the immune response during pathogenic virus have been previously described [35][36][37]. Currently, ferrets are used for influenza drug testing; for example, neuraminidase inhibitors are effective during ferret influenza infection [38][39][40].…”