The effect of antiapoptosis genes on clarification performance
The effect of antiapoptosis genes on clarification performance
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抗凋亡基因对澄清性能的影响
DOI：
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发表时间：
2014
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通讯作者：
A. Dileo
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作者：
Ajish S. R. Potty;A. Xenopoulos;Sonal Patel;H. Prentice;A. Dileo
最佳生物反应器收获时间通常基于最大化产物滴度而不损害产物质量来确定。我们建议在收获过程中还应考虑下游纯化的便利性。从这个角度，我们研究了抗凋亡基因对下游性能的影响。我们的假设是，更健壮的细胞将表现出更少的细胞裂解，从而产生更低水平的细胞碎片和宿主细胞污染物。我们重点关注澄清单元操作，测量澄清后浊度和宿主细胞蛋白（HCP）浓度作为生物反应器收获时间/细胞活力的函数。为了模拟使用叠盘离心的初步澄清，使用了由旋转盘（模拟离心机入口进料区的剪切力）和摆斗实验室离心机组成的缩小模型。我们的数据表明，在初步澄清（深层过滤器的典型）过程中不存在剪切的情况下，与对照细胞相比，观察到具有抗凋亡基因的细胞的HCP水平降低20-50%，离心后浊度降低50-65%。然而，在将细胞暴露于盘叠式离心机中典型的剪切水平时，HCP减少10-15%，而未观察到离心后浊度的差异。因此，使用涉及低剪切、<1 × 106 W/m3和对细胞损伤最小的澄清选项可实现抗凋亡基因的最大益处。© 2013美国化学工程师学会生物技术。程序，30：100-107，2014
Optimal bioreactor harvest time is typically determined based on maximizing product titer without compromising product quality. We suggest that ease of downstream purification should also be considered during harvest. In this view, we studied the effect of antiapoptosis genes on downstream performance. Our hypothesis was that more robust cells would exhibit less cell lysis and thus generate lower levels of cell debris and host‐cell contaminants. We focused on the clarification unit operation, measuring postclarification turbidity and host‐cell protein (HCP) concentration as a function of bioreactor harvest time/cell viability. In order to mimic primary clarification using disk‐stack centrifugation, a scale‐down model consisting of a rotating disk (to simulate shear in the inlet feed zone of the centrifuge) and a swinging‐bucket lab centrifuge was used. Our data suggest that in the absence of shear during primary clarification (typical of depth filters), a 20–50% reduction in HCP levels and 50–65% lower postcentrifugation turbidity was observed for cells with antiapoptosis genes compared to control cells. However, on exposing the cells to shear levels typical in a disk‐stack centrifuge, the reduction in HCP was 10–15% while no difference in postcentrifugation turbidity was observed. The maximum benefit of antiapoptosis genes is, therefore, realized using clarification options that involve low shear, <1 × 106 W/m3 and minimal damage to the cells. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 30:100–107, 2014