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<p>Thiols serve indispensable biochemical functions across catalysis, redox homeostasis and energy metabolism. However, profiling multiple thiols at the single-cell level remains challenging due to their trace amount and susceptibility to oxidation. Herein, we report an integrated strategy for thiol profiling at the single-cell level which combines live-cell labeling with organic mass cytometry. The live-cell labeling strategy facilitates the comprehensive measurement of intrinsic thiols with expanded coverage and improved sensitivity, while organic mass cytometry enables simultaneous quantification of 27 labeled thiols and 355 other metabolites from single cells. Assessment of metabolic fluctuation upon stimulation demonstrates practicability and accuracy of this integrated methodology which is capable of pathway activity monitoring, metabolic network mapping and untargeted metabolome profiling. Further application of this method in investigating RSL3-triggered ferroptosis reveals that RSL3 inhibits glutathione synthesis via nuclear factor E2-related factor 2- glutathione axis and results in heterogenous glutathione metabolism between subtypes.</p>

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Single-cell thiol profiling enabled by live-cell labeling reveals metabolic heterogeneity in ferroptosis

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年3月3日

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  • Daiyu Miao,
  • Qiuning Li,
  • Yi Zhang,
  • Shaojie Qin,
  • Ying Wang,
  • Xiaoyun Liu,
  • Yu Bai

摘要

Thiols serve indispensable biochemical functions across catalysis, redox homeostasis and energy metabolism. However, profiling multiple thiols at the single-cell level remains challenging due to their trace amount and susceptibility to oxidation. Herein, we report an integrated strategy for thiol profiling at the single-cell level which combines live-cell labeling with organic mass cytometry. The live-cell labeling strategy facilitates the comprehensive measurement of intrinsic thiols with expanded coverage and improved sensitivity, while organic mass cytometry enables simultaneous quantification of 27 labeled thiols and 355 other metabolites from single cells. Assessment of metabolic fluctuation upon stimulation demonstrates practicability and accuracy of this integrated methodology which is capable of pathway activity monitoring, metabolic network mapping and untargeted metabolome profiling. Further application of this method in investigating RSL3-triggered ferroptosis reveals that RSL3 inhibits glutathione synthesis via nuclear factor E2-related factor 2- glutathione axis and results in heterogenous glutathione metabolism between subtypes.