A Chemigenetic Fluorescence Lifetime Biosensor for Dopamine Sensing
- 1Department of Biomedical Engineering and Texas Materials Institute, The University of Texas at Austin, Austin, TX, USA
Abstract
Measuring the precise dynamics of specific neuromodulators in neurons is key to understanding how information is transmitted and processed in the brain. Despite the growing use of intensity-based neural activity sensors such as dLight in recent years, it remains challenging to monitor the dynamics of multiple neurochemicals simultaneously in live neurons. Unlike intensiometric sensors that are susceptible to artifacts caused by fluctuations in excitation power and rapid photobleaching, fluorescence lifetime-based sensors are less affected by these factors and can provide more stable measurements. Here we introduce dHaloLife635, a novel genetically encoded fluorescence-lifetime-based dopamine (DA) sensor that exhibits a fluorescence lifetime change up to -0.20 ns upon dopamine binding. dHaloLife635 consists of a G protein-coupled dopamine receptor (DRD1), a circularly permuted HaloTag (cpHaloTag), and a cell-permeable dye JF635 functionalized with a HaloTag ligand, a reactive chloroalkane linker (JF635-HTL). When dopamine binds to the dHaloLife635 sensor, it induces a conformational change in the cpHaloTag that alters the local environment of the conjugated dye, thereby changing its fluorescence lifetime. In live HEK293T cells and primary neurons, dHaloLife635 responds selectively to dopamine but not to serotonin, L-DOPA, or GABA, demonstrating excellent specificity. This dHaloLife635 sensor represents an initial step toward the establishment of a broader family of chemigenetic lifetime sensors that exhibit distinct fluorescence lifetime signatures upon binding their respective targets, thereby enabling highly specific and sensitive, real-time monitoring of multiple neurochemical dynamics in vivo.
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BibTeX
@misc{nguyen2026chemigenetic,
title = {{A Chemigenetic Fluorescence Lifetime Biosensor for Dopamine Sensing}},
author = {A.-T. Nguyen and H. Tran and L. Chen and A. Criswell and S. Bradbury and Y. Zhu and Y. He and S. Kim and Y.-I. Chen and T. D. Nguyen and S. Hong and Y.-A. Kuo and S. Seifi and W.-R. Chen and S. K. Seidlits and L. E. Fenno and H.-C. Yeh},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.19.752778},
url = {https://www.biorxiv.org/content/10.64898/2026.09.19.752778v1},
note = {CC BY-NC},
}
RIS
TY - UNPB
TI - A Chemigenetic Fluorescence Lifetime Biosensor for Dopamine Sensing
AU - A.-T. Nguyen
AU - H. Tran
AU - L. Chen
AU - A. Criswell
AU - S. Bradbury
AU - Y. Zhu
AU - Y. He
AU - S. Kim
AU - Y.-I. Chen
AU - T. D. Nguyen
AU - S. Hong
AU - Y.-A. Kuo
AU - S. Seifi
AU - W.-R. Chen
AU - S. K. Seidlits
AU - L. E. Fenno
AU - H.-C. Yeh
PY - 2026
DA - 2026/09/19
PB - bioRxiv
DO - 10.64898/2026.09.19.752778
UR - https://www.biorxiv.org/content/10.64898/2026.09.19.752778v1
AB - Measuring the precise dynamics of specific neuromodulators in neurons is key to understanding how information is transmitted and processed in the brain. Despite the growing use of intensity-based neural activity sensors such as dLight in recent years, it remains challenging to monitor the dynamics of multiple neurochemicals simultaneously in live neurons. Unlike intensiometric sensors that are susceptible to artifacts caused by fluctuations in excitation power and rapid photobleaching, fluorescence lifetime-based sensors are less affected by these factors and can provide more stable measurements. Here we introduce dHaloLife635, a novel genetically encoded fluorescence-lifetime-based dopamine (DA) sensor that exhibits a fluorescence lifetime change up to -0.20 ns upon dopamine binding. dHaloLife635 consists of a G protein-coupled dopamine receptor (DRD1), a circularly permuted HaloTag (cpHaloTag), and a cell-permeable dye JF635 functionalized with a HaloTag ligand, a reactive chloroalkane linker (JF635-HTL). When dopamine binds to the dHaloLife635 sensor, it induces a conformational change in the cpHaloTag that alters the local environment of the conjugated dye, thereby changing its fluorescence lifetime. In live HEK293T cells and primary neurons, dHaloLife635 responds selectively to dopamine but not to serotonin, L-DOPA, or GABA, demonstrating excellent specificity. This dHaloLife635 sensor represents an initial step toward the establishment of a broader family of chemigenetic lifetime sensors that exhibit distinct fluorescence lifetime signatures upon binding their respective targets, thereby enabling highly specific and sensitive, real-time monitoring of multiple neurochemical dynamics in vivo.
ER -