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High‐throughput metabolic profiling based on small amount of hepatic cells
- Zhou, Lina, Yin, Peiyuan, Luo, Ping, Tang, Ling, Wang, Zhichao, Gao, Peng, Piao, Hailong, Lu, Xin, Xu, Guowang
- Electrophoresis 2017 v.38 no.18 pp. 2296-2303
- amino acids, cell culture, detection limit, electrophoresis, fatty acids, human cell lines, liquid chromatography, mass spectrometry, metabolites, metabolomics
- Common metabolomics platforms require about 10⁶ cells, which has a limited throughput due to the time‐consuming steps of cell culture and preparation. There is a demand for metabolic profiling methods to improve analytical throughput and detection sensitivity based on small amount of cells. In this study, we proposed a high‐throughput scheme, integrating 96‐well plate cell cultivation, in‐situ cell pretreatment, and sensitive dansylation labeling coupled with LC‐MS analysis of metabolites inside HepG2 cells (of the order of magnitude of 10³ cells in each well). A simple and rapid cell pretreatment was performed showing good extraction efficiency and good precision (the RSDs smaller than 5%) for polar metabolites. The recovery in metabolite extraction evaluated with three isotope‐labeled amino acids was from 89.7 to 106.3% at low, medium, and high concentrations. The suitability of the method was illustrated by exploring influences of different fatty acids on HepG2 cells.