摘 要
本研究主要是探討利用新式生物反應器以固定化細胞培養技術生產日本腦炎病毒(Japanese encephalitis virus；JEV)之研究，首先根據所培養細胞的生長特性與原有生物反應器之缺點，利用已改良設計之新式生物反應器，評估此新式生物反應器適用性、效能，以及所培養細胞之生長代謝情形與病毒力價。整個實驗過程大致分為幾個階段，第一個階段探討細胞固定化培養之最適化培養條件與生長代謝情形，第二個階段找出細胞固定化培養於此新式生物反應器中最佳生長狀態，最後一個階段為病毒的培養。實驗後發現Vero細胞經固定化貼附於FIBRA-CEL®載體上，可擴大培養於新式生物反應器，Vero細胞最佳生長量達到6.6×106cells/mL。希望藉由此改良之新式生物反應器提供細胞與病毒一個良好之生長培養環境，獲得高產量、品質穩定一致之細胞生物製品，以提供ㄧ設備簡單與製程操作容易、低成本、低能源消耗之細胞製品生產基座。 / Abstract
In this study, we investigated the production of Japanese encephalitis virus (JEV) by the immobilized cell technology in a novel bioreactor. According to the disadvantages of original bioreactor and growth characteristics of cell culture, we evaluated the suitability and efficiency of a design-improved novel bioreactor as well as the growth and metabolic situation of cultured cells and titers of JEV. All studies including three major stages: (1) investigation of the optimal conditions and metabolic situation for the growth of immobilized cells, (2) finding the optimal conditions for the growth of immobilized cells in this novel bioreactor, and (3) growth of JEV using immobilized cells in this novel bioreactor. Our results showed that after immobilization on the FIBRA-CEL® carries, Vero cells can grow on the novel bioreactor up to the density of 6.6 × 106 cells/mL. Hopefully, the improvement of the novel bioreactor will provide an optimal growth condition for both the cells and viruses. Furthermore, it will also provide the basis for the production of cell products with advantages of simple-equipped, easy-to-operate, low cost, and low energy consumption. / 目 錄
誌謝------------------------------------------------- i
中文摘要 -------------------------------------------- ii
英文摘要 -------------------------------------------- iii
目錄 -------------------------------------------- iv
表目錄 -------------------------------------------- v
圖目錄 -------------------------------------------- vi
第一章 緒論---------------------------------------- 1
第二章 文獻探討------------------------------------ 3
第一節 日本腦炎病毒疫苗---------------------------- 3
第二節 動物細胞的培養------------------------------ 4
第三節 載體上動物細胞的培養------------------------ 5
第四節 動物細胞培養於生物反應器-------------------- 7
第三章 材料與方法---------------------------------- 10
一 細胞株的培養-------------------------------- 10
二 細胞冷凍保存與解凍培養---------------------- 10
三 細胞滾瓶培養-------------------------------- 11
四 病毒株的培養-------------------------------- 12
五 固定化載體材料製備-------------------------- 12
六 載體上細胞數的測定-------------------------- 12
七 細胞貼壁率的計算---------------------------- 13
八 生物反應器結構特性與固定化細胞培養---------- 13
九 日本腦炎病毒力價測定------------------------ 19
十 葡萄糖的測定-------------------------------- 19
第四章 結果與討論---------------------------------- 20
一 固定化載體材料比例對Vero細胞生長的影響------ 20
二 細胞貼附固定化時間對Vero細胞生長的影響------ 23
三 細胞接種量對Vero細胞生長的影響-------------- 24
四 生物反應器培養系統對Vero細胞生長的影響------ 25
五 新鮮培養基更換對Vero細胞生長的影響---------- 27
六 最適化細胞生長條件培養日本腦炎病毒---------- 29
第五章 結論與建議---------------------------------- 30
參考文獻 -------------------------------------------- 31
附錄一 PBS配製方法--------------------------------- 62
附錄二 Medium 199配製方法-------------------------- 62
附錄三 MEM medium配製方法-------------------------- 62
| Identifer | oai:union.ndltd.org:TW/094FY005108006 |
| Date | January 1994 |
| Creators | 王琪婷, Chi-ting Wang |
| Contributors | 廖明一, Ming-yi Liau |
| Publisher | 輔英科技大學, 生物技術系碩士班 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | 中文 |
| Detected Language | English |
| Type | 碩士 |
| Format | 63 |
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