RESEARCH ARTICLE Immunisation of Sheep with Bovine Viral Diarrhoea Virus, E2 Protein Using a Freeze- Dried Hollow Silica Mesoporous Nanoparticle Formulation

Bovine viral diarrhoea virus 1 (BVDV-1) is arguably the most important viral disease of cat-tle. It is associated with reproductive, respiratory and chronic diseases in cattle across the world. In this study we have investigated the capacity of the major immunological determi-nant of BVDV-1, the E2 protein combined with hollow type mesoporous silica nanoparticles with surface amino functionalisation (HMSA), to stimulate immune responses in sheep. The current work also investigated the immunogenicity of the E2 nanoformulation before and after freeze-drying processes. The optimal excipient formulation for freeze-drying of the E2 nanoformulation was determined to be 5 % trehalose and 1 % glycine. This excipient formu-lation preserved both the E2 protein integrity and HMSA particle structure. Sheep were immunised three times at three week intervals by subcutaneous injection with 500 μg E2 adsorbed to 6.2 mg HMSA as either a non-freeze-dried or freeze-dried nanoformulation. The capacity of both nanovaccine formulations to generate humoral (antibody) and cell-mediated responses in sheep were compared to the responses in sheep immunisation with Opti-E2 (500 μg) together with the conventional adjuvant Quil-A (1 mg), a saponin from th