Institute of Cell Culture Technology, Bielefeld University, Bielefeld, Germany.
Biotechnol Lett. 2011 Mar;33(3):495-502. doi: 10.1007/s10529-010-0466-7. Epub 2010 Nov 12.
A new method for sampling suspended animal cells by fast filtration is presented that allows rapid quenching of cellular metabolism and efficient separation of the cells from culture medium. Compared to sampling with a microstructure heat exchanger or centrifugation without prior quenching, the adenylate energy charge and the measured concentrations especially of metabolites with a high turnover rate or of metabolites early in metabolic pathways were substantially higher. No leakage of ATP from the cells was observed when using iso-osmotic NaCl solution in the washing step. The combination of fast filtration and cold methanol extraction is therefore suitable for intracellular metabolomic studies of suspended animal cell cultures and superior to other methods currently applied.
介绍了一种通过快速过滤来采样悬浮动物细胞的新方法,该方法可以快速淬灭细胞代谢,并有效地将细胞与培养基分离。与使用微结构热交换器或未经淬灭的离心法采样相比,三磷酸腺苷(adenylate energy charge)和特别是高周转率代谢物或代谢途径早期代谢物的测量浓度要高得多。在洗涤步骤中使用等渗氯化钠溶液时,没有观察到 ATP 从细胞中泄漏。因此,快速过滤和冷甲醇提取的组合非常适合悬浮动物细胞培养物的细胞内代谢组学研究,优于目前应用的其他方法。