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題名 | 利用震盪器培養家蠶細胞量產B型肝炎表面抗原蛋白之影響因素=Factors Affecting Mass Production of HBsAg in M-BmN Cells Cultured in Shaking Flasks |
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作者 | 王守良; 侯豐男; 王敏盈; | 書刊名 | 中華昆蟲 |
卷期 | 17:4 1997.12[民86.12] |
頁次 | 頁197-208 |
分類號 | 438.38 |
關鍵詞 | 昆蟲細胞培養; 震盪器; 饋料批次; B型肝炎表面抗原; Insect cell culture; Shaker; Fed-batch; Hepatitis B surface antigen; |
語文 | 中文(Chinese) |
中文摘要 | 本文主要探討M-BmN家蠶細胞株在懸浮狀況下培養於震盪器之情形,以了解有關攪拌速率、血清濃度、起始接種密度及操作體積等因子對細胞生長之影響。發現添加0.2%之Pluronic F-68於IPL-41培養基中,在高轉速(250 rpm)下,可有效地保護細胞生長於震盪瓶內,同時當利用饋料批次模式培養,在高轉速下,能提高最大細胞密度約30%(從32´l05至48´105 cells/ml)。在血清濃度方面,顯示M-BmN細胞在10%血清中的生長狀況比在5%胎牛血清為佳,尤其是利用饋料批次模式培養。而操作體積方面,則發現過大的體積(大於60ml)會導致溶氧量下降,使pH值偏低而不利於細胞生長。而在起始接種密度部份,發現較高的接種密度,細胞將生長更好,達到最大細胞密度的時間也較短。饋料批次培養模式不僅可增加細胞密度,亦可應用於增加異源重組蛋白之表現量,目前B型肝炎表面抗原之產量已可由批次培養所得之230 ng/ml,提高到利用饋料批次培養所得之570 ng/ml。 |
英文摘要 | This study describes the effect of stirring speed, concentration of serum, working volume and mode of culture methods on cell growth of the M-BmN cell linewhich was established from Bombyx mori. The cells could be cultured in suspensionin shaking flasks at a speed of 80 rpm or below. If 0.2% of Pluronic F-68 was added into the IPL-41 medium, the cells could grow, even if the stirring speed was increased to 250 rpm. Studies on the effect of fetal bovine serum (FBS) concentrations on cell growth show that cells grew better in medium supplemented with 10% FBS than did those with 5% FBS. We also found that, when the working volume was 60 ml or more in a 250ml shaking flask, the pH of the culture medium kept dropping and finally inhibited cell growth. In contrast, change of pH values inthe culture medium, for cells growing in a flask with a working volume of 30ml, first dropped, but then increased again. The increase of pH was used as a sign to indicate the timing to feed glucose and glutamine in a fed-batch culture method. This method, when applied to MBm-N cells culture, can increase the maximum celldensity from 32×10□ to 48×10□ cells/ml. More significantly, the production of HBsAg could also be increased from 230 to 560 ng/ml. |
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