Dalm Marcella C F, Jansen Menno, Keijzer Timo M P, van Grunsven Wout M J, Oudshoorn Arthur, Tramper Johannes, Martens Dirk E
Department of Agrotechnology and Food Sciences, Food and Bioprocess Engineering Group, Wageningen University, Bomenweg 2, 6703 HD Wageningen, The Netherlands.
Biotechnol Bioeng. 2005 Sep 30;91(7):894-900. doi: 10.1002/bit.20552.
Perfusion systems have the possibility to be operated continuously for several months. It is important that the performance of the cell retention device does not limit the operation time of a perfusion process used in the production of active pharmaceutical ingredients. Therefore, the aim of this study was to investigate the reliability and long-term stability of an acoustic perfusion process using the 200 L/d BioSep. As the BioSep is an external device, it is possible that dependent on the recirculation rate nutrient gradients occur in the external loop, which could affect the cell metabolism. Therefore, the effect of possible nutrient gradients on cell metabolism, viability and productivity was studied by varying the recirculation rate. In this study, it is shown that a perfusion process using a pilot-scale acoustic cell-retention device (200 L/d) is reliable and simple to operate, resulting in a stable 75-day cultivation of a hybridoma cell line producing a monoclonal antibody. The recirculation rate had a significant effect on the oxygen concentration in the external loop, with oxygen being depleted within the cell-retention device at recirculation rates below 6 m3/m(reactor)3.d (=600 L/d). The oxygen depletion at low circulation rates correlated with a slightly increased lactate production rate. For all other parameters no effect of the recirculation rate was observed, including cell death measured through the release of lactate dehydrogenase and specific productivity. A maximum specific productivity of 12 pg/cell.d was reached.
灌注系统有可能连续运行数月。细胞截留装置的性能不限制用于生产活性药物成分的灌注过程的运行时间，这一点很重要。因此，本研究的目的是使用200 L/d的BioSep研究声学灌注过程的可靠性和长期稳定性。由于BioSep是一个外部装置，取决于再循环速率，外部回路中可能会出现营养物质梯度，这可能会影响细胞代谢。因此，通过改变再循环速率，研究了可能的营养物质梯度对细胞代谢、活力和生产力的影响。在本研究中，结果表明，使用中试规模的声学细胞截留装置（200 L/d）的灌注过程可靠且操作简单，可使产生单克隆抗体的杂交瘤细胞系稳定培养75天。再循环速率对外部回路中的氧气浓度有显著影响，当再循环速率低于6 m³/（反应器）³·d（=600 L/d）时，细胞截留装置内的氧气会耗尽。低循环速率下的氧气耗尽与乳酸产生速率略有增加相关。对于所有其他参数，未观察到再循环速率的影响，包括通过乳酸脱氢酶释放测量的细胞死亡和比生产力。最大比生产力达到12 pg/细胞·d。
