| Summary: | 碩士 === 國立屏東科技大學 === 動物疫苗科技研究所 === 105 === Flagellin from Salmonella is a highly conserved protein that elicits TLR5-dependent inflammatory responses and plays an important role. In the literature, The TLR-5 recognition site are 80~119、404~433and 420~451 for flagellin amino acid sequence, so we use FliC as the goal of cloning. Chitosan is similar to cellulose, by chitin through deacetylation, to remove part or all of the acetyl group and exposed from the free amino groups.It can elicit immune response, anti-tumor and anti-microbial infection. In recent years, Chitosan was treated as adjuvant. In addition also found that chitosan has antibacterial effect, can increase immune for Candida albicans and to kill the Candida. In nature, not only chitosan has the effect of promoting immune response, For example: Glucan and mannose also found to have anti-microbial and anti-bacterial infection protection. In recent years, the literature confirmed that chitosan modified mannose ligand, can improve macrophage ,dendritic cells and other phagocytic cells of antigen-presenting cell and mannose receptors, to improve the vaccine immunogenicity. Not only that, mannosylated chitosan (MC) has a longer release time than chitosan.
In our study, we evaluated the adjuvant effect of flagellin and mannosylated chitosan. We already cloned flagellin and ligated to pET32a vector for transformation and expression in E. coli BL21.SDS-PAGE and Western blot analysis showed that the protein is 56KDa .
Then the expressed flagellin was combined with the bovine serum albumin(BSA) and mannosylated chitosan and we vaccinated 6week-old mouse, and evaluated using ELISA ,cytokine expression analysis, and lymphocyte proliferation assay. Result shows that group of BSA combined flagellin and mannosylated chitosan,the IgG antibody and T cell proliferation other groups.Part of cytokine expression analysis, group of BSA combined mannosylated chitosan can elicits IL-6.
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