Liao Ning virus (LNV) is an emerging arbovirus first isolated in 1999 from mosquitoes in China. The virus readily replicates in mammalian cells and is known to cause lethal haemorrhagic symptoms in laboratory-infected mice. In 2013, LNV was first detected in mosquito species in Australia (Coffey et al., 2014). We have subsequently isolated LNV from four different genera of mosquitoes (Culex, Anopheles, Mansonia and Aedes) in New South Wales, The Northern Territory, Queensland and Western Australia; collected between 2000 and 2014. The evidence suggests that multiple vertebrate hosts could be involved in the natural transmission cycle of LNV. However, limited research tools have prevented investigation into the epidemiology and potential medical and veterinary significance of the virus. We have purified LNV from an infected mosquito sample and prepared antisera to use as a positive reference control for serological analysis of LNV. Furthermore, we have produced and characterised the first LNV-specific monoclonal antibody (6E6). Collectively these tools have been used to develop two serological assays (fixed-cell ELISA and micro-neutralisation assay) for the detection of LNV-specific antibodies in serum from Australian vertebrates. Preliminary results identified rabbits and humans as potential hosts, representing the first serological evidence of natural infections of a vertebrate with LNV.