Reuveny S, Velez D, Macmillan J D, Miller L
J Immunol Methods. 1986 Jan 22;86(1):53-9. doi: 10.1016/0022-1759(86)90264-4.
Environmental and cultural factors that could affect growth and cell viability of mouse-mouse hybridoma cells in culture were investigated. The aim was to determine conditions that could prolong viability and result in increased yields of monoclonal antibodies in stirred reactors. Factors studied included temperature, level of dissolved oxygen, nutrient depletion, and waste product accumulation. Growing cells at temperatures 3-9 degrees lower than optimum (37 degrees C) increased viability but monoclonal antibody production was lowered. A low level of dissolved oxygen (25% air saturation compared to 60% for optimum growth) prolonged cell viability and increased the monoclonal antibody yield by about 50%. Feeding cultures daily to maintain the glucose level above 1 mg/ml and at the same time feeding cells glutamine (150 micrograms/10(6) cells per day) maintained the level of viable cells at 1.7 X 10(6)/ml for at least 9 days and resulted in an antibody yield of 290 micrograms/ml, about a 70% increase over cultures fed either glucose or glutamine alone. Ammonium ion, added to cell populations at levels produced in cultures, stopped cell growth and decreased antibody production. Another waste product, lactic acid, had no toxic effect when added to media at levels found in cultures. These results agree with our suggestion that monoclonal antibody production is enhanced by maintaining cell viability over a prolonged period and provide a base for investigating modes of hybridoma cell propagation in fermentors.
研究了可能影响培养中的小鼠 - 小鼠杂交瘤细胞生长和细胞活力的环境和文化因素。目的是确定能够延长活力并导致搅拌反应器中单克隆抗体产量增加的条件。研究的因素包括温度、溶解氧水平、营养物质消耗和废物积累。在比最佳温度（37℃）低3 - 9℃的温度下培养细胞可提高活力，但单克隆抗体产量会降低。低溶解氧水平（25%空气饱和度，最佳生长为60%）可延长细胞活力，并使单克隆抗体产量提高约50%。每天向培养物中添加以维持葡萄糖水平高于1mg/ml，同时向细胞中添加谷氨酰胺（每天150微克/10⁶个细胞），可使活细胞水平维持在1.7×10⁶/ml至少9天，并产生290微克/ml的抗体产量，比单独添加葡萄糖或谷氨酰胺的培养物增加约70%。以培养物中产生的水平向细胞群体中添加铵离子会停止细胞生长并降低抗体产量。另一种废物乳酸，以培养物中发现的水平添加到培养基中时没有毒性作用。这些结果与我们的建议一致，即通过长时间维持细胞活力可提高单克隆抗体产量，并为研究杂交瘤细胞在发酵罐中的增殖模式提供了基础。
