Single-Cell Mass Spectrometry of Metabolites Extracted from Live Cells by Fluidic Force Microscopy
Single-Cell Mass Spectrometry of Metabolites Extracted from Live Cells by Fluidic Force Microscopy
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DOI：
10.1021/acs.analchem.7b00367
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发表时间：
2017-05-02
影响因子：
7.4
通讯作者：
Vorholt, Julia A.
中科院分区：
文献类型：
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作者：
Guillaume-Gentil, Orane;Rey, Timo;Vorholt, Julia A.
单细胞代谢物分析提供了有关细胞功能和对外部刺激反应的有价值的信息。虽然质谱法的最新进展达到了研究单细胞中代谢物所需的灵敏度，但目前的方法通常会分离和牺牲细胞，造成干扰状态并阻止互补分析。在这里，我们提出了一个两步的方法，结合使用流体力显微镜，其次是基质辅助激光解吸/电离飞行时间质谱的亚皮升分辨率的细胞内液的无损和定量撤出。所开发的方法能够检测和鉴定从单个HeLa细胞的细胞质中回收的20种代谢物。该方法在C-13-葡萄糖喂养实验中得到进一步验证，该实验显示标记的碳原子掺入不同的代谢物中。代谢物取样，然后进行质谱测量，能够保存所分析细胞的生理环境和活力，为代谢物分析之前、期间和之后的细胞补充分析提供机会
Single-cell metabolite analysis provides valuable information on cellular function and response to external stimuli. While recent advances in mass spectrometry reached the sensitivity required to investigate metabolites in single cells, current methods commonly isolate and sacrifice cells, inflicting a perturbed state and preventing complementary analyses. Here, we propose a two-step approach that combines nondestructive and quantitative withdrawal of intracellular fluid with subpicoliter resolution using fluidic force microscopy, followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The developed method enabled the detection and identification of 20 metabolites recovered from the cytoplasm of individual HeLa cells. The approach was further validated in C-13-glucose feeding experiments, which showed incorporation of labeled carbon atoms into different metabolites. Metabolite sampling, followed by mass spectrometry measurements, enabled the preservation of the physiological context and the viability of the analyzed cell, providing opportunities for complementary analyses of the cell before, during, and after metabolite analysis