| Summary: | 碩士 === 國立成功大學 === 生物化學暨分子生物學研究所 === 102 === Dogs are the most faithful friends for people. The statistics shows that every four Americans raise a dog. Every year the medical expense for canine diseases was 10 billion USD. Our previous studies showed that the IL-20 monoclonal antibody have protective function in osteoporosis and breast cancer. These diseases also occur in the dog. Over the past decade, cancer is the leading cause of death in older dogs. Breast cancer is the first leading cause of death in old female dogs. Thus, there is significant demand for new drug to treat canine cancer. Therefore, I was aimed to generate a neutralizing anti-IL-20 monoclonal antibody against dog. We expressed and purified the recombinant canine IL-20 protein from mammalian cells and E. coli cells. IL-20 protein and the adjuvant mixture used as an antigen was injected into mice to generate the anti-canine IL-20 antibody. I successfully established three stable hybridoma clones. ELISA, western blotting and IHC staining showed that each monoclonal antibody specifically recognized canine IL-20. They also cross-reacted with mouse and rat IL-20. In these three clones, 8H has the highest binding affinity to canine IL-20. Here I select 8H antibody for further characterization and functional analysis. It is immunoglobulin gamma 1 (mIgG1) subtype. I also mapped the epitope of canine IL-20 recognized by the 8H antibody using western blotting. I have sequenced the variable region of 8H antibody by 5’RACE. The preliminary result showed that the 8H antibody inhibited canine and mouse breast cancer cell migration in vitro and in vivo. In the future, I will construct 8H canine chimeric antibody and investigate its therapeutic potential in clinics
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