In HCV infection, it has been noted that despite hundreds to thousands of viral variants in the source, only 1-3 ‘founders’ establish infection, with a second strong bottleneck occurring at ~100 days post infection. Analysis of the envelope (E)1 and E2 regions revealed that at this second bottleneck all founder variants were extinguished - irrespective of disease outcome. In chronic progressors new variants emerged that carried mutations within the E region. Thus, it is likely that rapid evolution within E1/E2 allows variants to escape neutralisation and establish persistent infection. In the present study, we aim to use autologous and heterologous Envelope viruses to identify whether timing and breadth of the neutralising antibody (nAb) response contribute to the emergence of escape variants and the establishment of persistent infection. HCV seronegative injecting drug users are followed for incident infection as assessed by HCV RNA positivity. Blood samples for incident cases (n=15) are collected fortnightly over 24 weeks before viral clearance (n=7) or chronicity (n=8) is resolved. Near-full length HCV genome next generation sequencing (NGS) is utilised to examine the founder sequence and chronic variants. E1/E2 regions, representing the viral populations of interest, are cloned into expression plasmids and HCV pseudoparticles (HCVpp) are produced. Neutralisation activity is determined against autologous HCVpp and a library of heterologous HCVpp representing GT1-6 to determine timing and breadth of the nAb responses in patient sera respectively. To date, 15 incident subjects with longitudinal sampling have been selected. NGS sequence analysis has been performed longitudinally on all subjects. Autologous HCVpp representing founder and chronic variants, and heterologous HCVpp from GT1-6 have been constructed. Longitudinal nAb responses targeting autologous founder virus have been measured in clearer subject, Cl_168. The nAb responses, although weak, were detectable from 5DPI and gradually increased before peaking at ~58DPI and then persisted after clearance. Peak in nAb correlated with a decrease in viral load and subsequent clearance suggesting nAbs contribute toward clearance of the founder virus. We hypothesise that the emergence of nAb responses will be early in both clearers and chronic progressors but rapidly broaden in breadth in clearers. In this study viral variants associated with transmission and persistent infection in early primary infection have been synthesised and the autologous responses against these variants is underway.