Wije Munige Shakya, Bhusal Deepti, Peng Zongkai, Chen Dan, Yang Zhibo
Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma 73019, United States.
Stephenson Cancer Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104, United States.
JACS Au. 2025 May 6;5(5):2379-2387. doi: 10.1021/jacsau.5c00327. eCollection 2025 May 26.
Single-cell mass spectrometry (SCMS) has emerged as a powerful tool for analyzing metabolites in individual cells, including live cells. However, cell metabolites have a rapid turnover rate, whereas maintaining metabolites' profiles of live cells during sample transport, storage, or extended measurements can be challenging. In this study, a cell preparation method, which integrates cell washing by volatile salt solution, rapid liquid nitrogen (LN) quenching, freeze-drying in vacuum, and freezer storage at -80 °C, to preserve cell metabolites for SCMS measurement is discussed. Experimental results revealed that LN quenching preserved the overall cell metabolome, whereas storage at -80 °C for 48 h slightly changed the metabolite profiles in quenched cells. However, metabolites in unquenched cells were changed regardless of low-temperature storage. The influence of omission of quenching and low-temperature storage on cell metabolites and relevant pathways were investigated. Results from this work indicate that cell quenching is necessary, but low-temperature storage time should be minimized to preserve cell metabolites. The method developed in the current work can be readily adopted by SCMS techniques with storage remaining largely unaltered, allowing for extended SCMS studies.
单细胞质谱分析（SCMS）已成为分析单个细胞（包括活细胞）中代谢物的强大工具。然而，细胞代谢物的周转速度很快，而在样品运输、储存或长时间测量过程中维持活细胞的代谢物谱具有挑战性。在本研究中，讨论了一种细胞制备方法，该方法将挥发性盐溶液洗涤细胞、快速液氮（LN）淬灭、真空冷冻干燥以及在-80°C冷冻保存相结合，以保存细胞代谢物用于SCMS测量。实验结果表明，液氮淬灭保留了整体细胞代谢组，而在-80°C储存48小时会使淬灭细胞中的代谢物谱略有变化。然而，未淬灭细胞中的代谢物无论低温储存与否都会发生变化。研究了省略淬灭和低温储存对细胞代谢物及相关途径的影响。这项工作的结果表明，细胞淬灭是必要的，但低温储存时间应尽量缩短以保存细胞代谢物。当前工作中开发的方法可被SCMS技术轻松采用，且储存情况基本保持不变，从而能够进行扩展的SCMS研究。
