High-Sensitivity, Dual Ratiometric and Turn-on ABATaRs for Multiplexed Temporal Imaging of Ferroptosis Hallmarks in Living Cells
5 affiliations
- 1Department of Chemical Sciences
- 2Tata Institute of Fundamental Research
- 3Department of Applied Physics
- 4University of Osaka
- 5Institute for Open and Transdisciplinary Research Initiatives
Abstract
We present a computationally-guided workflow for designing dual turn-on and ratiometric Raman sensors. The workflow has been used to rationally design activity-based alkyne tagged Raman, ABATaR , sensors that overcome the inherently low Raman sensitivity via strategic placement of electron-donating and electron-withdrawing groups. An ABATaR sensing scaffold designed via the workflow afforded ~ 9000 times higher Raman scattering cross-section than the dimethylsulfoxide C-H stretching-mode. The sensitivity of the scaffold was compared to that of 5-ethynyl-2'-deoxyuridine ( EdU ) to determine Raman intensity versus EdU , RIE, a parameter used to benchmark Raman probes. An RIE value of 170 was obtained which is one of the highest amongst previously reported monoyne and diyne-based Raman probes. The sensitive ABATaR -scaffold was developed into a dual turn-on and ratiometric Fe 2+ -sensor that was applied to distinctly quantify endogenous cellular labile Fe 2+ ion levels. In a key result, live-cell, temporal imaging with the Fe 2+ - ABATaR sensor revealed that intracellular labile Fe 2+ levels progressively increase with ferroptosis while the total Fe-level remains constant. To demonstrate the generalizability of the design workflow, we synthesized an orthogonal ABATaR -sensor for H 2 O 2 . The H 2 O 2 -sensor was applied together with the Fe 2+ -sensor along with inherent Raman peaks for cytochromes to enable simultaneous, multiplexed spontaneous Raman imaging of three distinct ferroptosis hallmarks—labile Fe 2+ , H 2 O 2 , and redox-sensitive cytochrome Raman signals—in living cells. Importantly, the ability to pre-design orthogonal ratiometric Raman sensors and simultaneously track three regulators of ferroptosis via discrete non-overlapping Raman peaks highlights the tremendous potential of Raman sensors toward attaining live cell multiplexed imaging and quantification.
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BibTeX
@misc{das2026highsensitivity,
title = {{High-Sensitivity, Dual Ratiometric and Turn-on ABATaRs for Multiplexed Temporal Imaging of Ferroptosis Hallmarks in Living Cells}},
author = {Sujit K Das and Heqi Xi and Saurabh Kumar and Katsumasa Fujita and Ankona Datta},
howpublished = {ChemRxiv},
year = {2026},
month = oct,
doi = {10.26434/chemrxiv.15010169/v1},
url = {https://chemrxiv.org/doi/full/10.26434/chemrxiv.15010169/v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - High-Sensitivity, Dual Ratiometric and Turn-on ABATaRs for Multiplexed Temporal Imaging of Ferroptosis Hallmarks in Living Cells
AU - Sujit K Das
AU - Heqi Xi
AU - Saurabh Kumar
AU - Katsumasa Fujita
AU - Ankona Datta
PY - 2026
DA - 2026/10/09
PB - ChemRxiv
DO - 10.26434/chemrxiv.15010169/v1
UR - https://chemrxiv.org/doi/full/10.26434/chemrxiv.15010169/v1
AB - We present a computationally-guided workflow for designing dual turn-on and ratiometric Raman sensors. The workflow has been used to rationally design activity-based alkyne tagged Raman, ABATaR , sensors that overcome the inherently low Raman sensitivity via strategic placement of electron-donating and electron-withdrawing groups. An ABATaR sensing scaffold designed via the workflow afforded ~ 9000 times higher Raman scattering cross-section than the dimethylsulfoxide C-H stretching-mode. The sensitivity of the scaffold was compared to that of 5-ethynyl-2'-deoxyuridine ( EdU ) to determine Raman intensity versus EdU , RIE, a parameter used to benchmark Raman probes. An RIE value of 170 was obtained which is one of the highest amongst previously reported monoyne and diyne-based Raman probes. The sensitive ABATaR -scaffold was developed into a dual turn-on and ratiometric Fe 2+ -sensor that was applied to distinctly quantify endogenous cellular labile Fe 2+ ion levels. In a key result, live-cell, temporal imaging with the Fe 2+ - ABATaR sensor revealed that intracellular labile Fe 2+ levels progressively increase with ferroptosis while the total Fe-level remains constant. To demonstrate the generalizability of the design workflow, we synthesized an orthogonal ABATaR -sensor for H 2 O 2 . The H 2 O 2 -sensor was applied together with the Fe 2+ -sensor along with inherent Raman peaks for cytochromes to enable simultaneous, multiplexed spontaneous Raman imaging of three distinct ferroptosis hallmarks—labile Fe 2+ , H 2 O 2 , and redox-sensitive cytochrome Raman signals—in living cells. Importantly, the ability to pre-design orthogonal ratiometric Raman sensors and simultaneously track three regulators of ferroptosis via discrete non-overlapping Raman peaks highlights the tremendous potential of Raman sensors toward attaining live cell multiplexed imaging and quantification.
ER -