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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.