Extraction of Aqueous Metabolites from Cultured Adherent Cells for Metabolomic Analysis by Capillary Electrophoresis-Mass Spectrometry
Extraction of Aqueous Metabolites from Cultured Adherent Cells for Metabolomic Analysis by Capillary Electrophoresis-Mass Spectrometry
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DOI：
10.3791/59551
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发表时间：
2019-06-01
影响因子：
1.2
通讯作者：
Makinoshima, Hideki
中科院分区：
文献类型：
--
作者：
Maruyama, Ami;Kami, Kenjiro;Makinoshima, Hideki
代谢组学分析是一种很有前途的组学方法，不仅可以了解癌细胞与正常细胞相比的特定代谢调控，还可以识别用于早期癌症检测和预测癌症患者化疗反应的生物标志物。制备用于代谢组学分析的均匀样品是一个仍有待解决的关键问题。在这里，我们提出了一个简单而可靠的协议，从培养的贴壁细胞中提取代谢物的代谢组学分析，使用毛细管电泳-质谱（CEMS）。通过培养和洗涤细胞、用甲醇处理细胞、提取代谢物以及用离心柱去除蛋白质和大分子用于CE-MS分析来分析来自培养细胞的水性代谢物。使用二酰胺（一种氧化试剂）处理的肺癌细胞系的代表性结果说明了在氧化应激下细胞的明显可观察到的代谢转变。这篇文章将是特别有价值的学生和研究人员参与代谢组学研究，谁是新的收获代谢产物从细胞系分析CE-MS。
Metabolomic analysis is a promising omics approach to not only understand the specific metabolic regulation in cancer cells compared to normal cells but also to identify biomarkers for early-stage cancer detection and prediction of chemotherapy response in cancer patients. Preparation of uniform samples for metabolomic analysis is a critical issue that remains to be addressed. Here, we present an easy and reliable protocol for extracting aqueous metabolites from cultured adherent cells for metabolomic analysis using capillary electrophoresis-mass spectrometry (CEMS). Aqueous metabolites from cultured cells are analyzed by culturing and washing cells, treating cells with methanol, extracting metabolites, and removing proteins and macromolecules with spin columns for CE-MS analysis. Representative results using lung cancer cell lines treated with diamide, an oxidative reagent, illustrate the clearly observable metabolic shift of cells under oxidative stress. This article would be especially valuable to students and investigators involved in metabolomics research, who are new to harvesting metabolites from cell lines for analysis by CE-MS.