Selection of <it>Neospora caninum </it>antigens stimulating bovine CD4<sup>+ve </sup>T cell responses through immuno-potency screening and proteomic approaches
Rocchi Mara S | Bartley Paul M | Inglis Neil F | Collantes-Fernandez Esther | Entrican Gary | Katzer Frank | Innes Elisabeth A
<p>Abstract</p> <p><it>Neospora caninum </it>is recognised worldwide as a major cause of bovine infectious abortion. There is a real need to develop effective strategies to control infection during pregnancy which may lead to either abortion or congenital transmission. Due to the intracellular nature of the parasite, cell-mediated immune (CMI) responses involving CD4<sup>+ve</sup>, CD8<sup>+ve</sup>, γ/δ TCR<sup>+ve </sup>T cells and NK cells, as well as production of IFN-γ, are thought to be important for protective immunity. In this study we applied a combination of proteomic and immunological approaches to identify antigens of <it>N. caninum </it>that are recognized by CD4<sup>+ve </sup>T cell lines derived from infected cattle. Initially, <it>N. caninum </it>tachyzoite Water Soluble Antigens (NcWSA) were fractionated by size-exclusion HPLC and then screened for immune-potency using CD4<sup>+ve </sup>T cell lines. LC-ESI-MS/MS (liquid chromatography electrospray ionisation tandem mass spectrometry) was employed to catalogue and identify the proteins comprising three immunologically selected fractions and led to the identification of six <it>N. caninum </it>target proteins as well as sixteen functional orthologues of <it>Toxoplasma gondii</it>. This approach allows the screening of biologically reactive antigenic fractions by the immune cells responsible for protection (such as bovine CD4<sup>+ve </sup>cells) and the subsequent identification of the stimulating components using tandem mass spectrometry.</p>
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