Development of a highly sensitive serological assay for Covid-19 based on the use of virus-like particles and protein-based nanoparticles.

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We propose to use the baculovirus expression system to generate Virus-Like Particles (VLPs) mimicking the recently emergent SARS-CoV-2. We have previously co-expressed three essential structural proteins (S, M, and E) of SARS-CoV using a single baculovirus expression vector developed in our lab and demonstrated that when co-expressed in Sf9 cells, these three proteins were able to spontaneously adopt a conformation mimicking the full virus (1). Subsequent mouse studies showed that vaccination with these elicited a strong neutralising antibody response, equivalent to that seen in a convalescent human. Recently we used these pre-existing reagents as a basis to produce SARS-CoV-2 VLPs, substituting the S, M, and E proteins into the previously generated Baculovirus constructs, and preliminary data shows that all three SARS-CoV-2 proteins are expressed. We propose to characterise thoroughly the expressed proteins, the formation of VLPs, and the immune response in mice to validate their authenticity as virus mimics. VLPs will be ultimately used to develop an antibody detection assay. In parallel, we will express the SARS-CoV-2 Fc-tagged S protein alone to be externally encapsulated with the lumazine synthase protein-based nanoparticles (PNPs) that we have already prepared. We have shown that similar nanoparticles for a receptor binding protein of a different virus enhanced binding affinity and avidity. These PNPs self-assemble into hollow monodisperse spherical structures, which stabilises the trimeric S protein and enhances antigenicity (2). Both SARS-CoV-2 VLPs and PNPs are excellent candidates for the development of highly sensitive diagnostic assay that will be used to detect COVID-19 infected human sera.