Of the arboviruses that are NIAID Category A, B and C Priority Pathogens, approved vaccines are available only for Japanese encephalitis virus, tick-borne encephalitis virus, and yellow fever virus (YFV). Additionally, use of even the highly effective YFV17D vaccine is now being re-evaluated due to increasing reports of severe adverse events following vaccination, particularly in immunocompromised patients. There are currently no other prophylactic or therapeutic strategies approved for arbovirus diseases. Thus, we propose to investigate the utility of human (Hu) and humanized murine (MuHu) arbovirus-specific monoclonal antibodies (MAbs) as antiviral prophylactic or therapeutic reagents. Initial studies, including those in our Progress Report below, indicate that MAbs can be effective prophylactic and therapeutic agents for the encephalitic West Nile and Venezuelan equine encephalitis viruses (VEEV) in mouse models of infection. Based on these results we hypothesize that antiviral MAb prophylaxis and/or therapy using Hu- or MuHuMAbs will also be successful for the viscerotropic, Category A and C arboviruses dengue (DENV) and yellow fever (YFV). We will continue to assess the protective and therapeutic capacities of HuMAb for VEEV developed during our first grant period and will use two approaches to develop YFV and DENV2-reactive HuMAb or MuHuMAb. 1) Use new cell fusion partners, MFP2 or MFP2D cells, to prepare human hybridomas using either human peripheral blood B-cells from infection-immune persons, or human B-cells from virus-immunized, humanized mice. 2) Genetically recombine the variable regions of protective murine MAbs with the constant regions of human immunoglobulin. Protective and therapeutic capacity of these MAbs will be tested by peripheral virus challenge of outbred mice (VEEV) or inbred mice deficient in their interferon response (YFV and DENV2). Specific aims are as follows: 1. Continue characterization and assessment of protective and therapeutic capacities of human or humanized VEEV-reactive MAbs and map the human anti-VEEV repertoire. 2. Develop HuMAbs and MuHuMAbs specific for YFV and DENV2 Eproteins. 3. Develop a new peripheral challenge animal model for YF using YFV17D vaccine in interferondeficient AG129 mice. 4. Characterize the ability of the YFV- and DENV2-reactive MuHuMAbs and HuMAbs to protect from or treat YFV and DENV2 infections in the AG129 mouse model and use the HuMAbs to map the human anti-E protein antibody repertoire. This project fits within the RMRCE Integrated Research Focus on Viral Therapeutics and will interact directly with RPs 3.1 and 1.7 and Core C.