Toolkit for the study of yellow fever virus

Abstract

Yellow fever virus (YFV) is a dangerous re-emerging mosquito borne flavivirus with a high lethality, which causes untreatable haemorrhagic fever in humans. Important disadvantages regarding safety and limited vaccine supply have been identified in the existing live attenuated vaccine. Alternative safer therapeutic strategies are needed to reduce the lethality of infections and protect a wider group of people. To this point, we created an array of essential tools for the study of YFV. We designed a novel YFV reporter virus to develop a highly sensitive high throughput neutralisation assay (Z’=0.65), which can screen virus inhibitors in only 48 hours. Moreover, we designed a site-specific biotinylated soluble YF envelope (E) protein by a cloning strategy involving the use of a biotin acceptor peptide and E. coli Biotin-protein birA ligase. The biotinylated E protein was used to develop a highly sensitive ELISA to screen antibodies that bind to the YFV E protein. Furthermore, we immunised mice with a soluble YFV E protein and generated anti-YFV E IgG secreting hybridoma cell lines. Monoclonal antibodies secreted by hybridomas were purified by affinity chromatography and are being characterised. The developed antibodies against YF E protein are a crucial tool for the molecular study of YF. Additionally, neutralising antibodies against YFV could potentially be developed into the first therapeutic treatment against YF infection. The toolkit developed in this project includes a neutralisation assay, a binding ELISA, and anti YFV E antibodies. These are essential instruments to expand the knowledge of YFV.