Evaluation of cell damage caused by cold sampling and quenching for metabolome analysis
Evaluation of cell damage caused by cold sampling and quenching for metabolome analysis
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DOI：
10.1007/s00253-011-3377-1
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发表时间：
2011-12-01
影响因子：
5
通讯作者：
Franco-Lara, Ezequiel
中科院分区：
文献类型：
--
作者：
Schaedel, Friederike;David, Florian;Franco-Lara, Ezequiel
在整个样品水平上使用基于OD的方法和ATP损失研究以及在单细胞水平上通过流式细胞术研究了用于代谢组学分析的采样和淬灭期间的细胞损伤。大肠杆菌在摇瓶中培养，并在指数生长期期间取样到几种冷淬灭溶液中，改变淬灭溶液组成和取样温度。对于单细胞分析，将样品与选择性碘化丙啶染料一起孵育，并通过流式细胞术进行分析以区分完整细胞和受损细胞。结果表明，淬火溶液、温度或冷却速率的每一种组合都会影响E。大肠杆菌细胞膜完整性表明破裂，这不仅会让染料进入，而且会让细胞内ATP离开细胞，这在体内代谢组学分析中是不希望的。
Cell damage during sampling and quenching for metabolome analysis have been investigated at whole sample level using an OD-based method and ATP loss investigation, and at single cell level by means of flow cytometry. Escherichia coli was cultivated in shake flasks and sampled into several cold quenching solutions during exponential growth phase varying quenching solution composition and sampling temperature. For single cell analysis, the samples were incubated with selective propidium iodide dye and analysed via flow cytometry to differentiate between intact and damaged cells. It was found that every combination of quenching solution, temperature, or cooling rate tested influenced the E. coli cell membrane integrity indicating rupture which will not only let the dye in, but also intracellular ATP out of the cells, which is not desired in in vivo metabolome analysis.