These are preliminary reports that have not been peer-reviewed. They should not be regarded as conclusive, guide clinical practice/health-related behavior, or be reported in news media as established information. For more information, please see our FAQs.
2 files

Fluorescence lifetime predicts performance of voltage sensitive fluorophores in cardiomyocytes and neurons

revised on 23.06.2020, 19:42 and posted on 25.06.2020, 10:11 by Steven Boggess, Julia Lazzari-Dean, Benjamin Raliski, Dong Min Mun, Amy Li, Evan Miller

Voltage imaging with fluorescent indicators offers a powerful complement to traditional electrode or Ca2+-imaging approaches for monitoring electrical activity. Small molecule fluorescent indicators present the unique opportunity for exquisite control over molecular structure, enabling detailed investigations of structure/function relationships. In this paper, we tune the conjugation between aniline donors and aromatic π systems within the context of photoinduced electron transfer (PeT) based voltage indicators. We describe the design and synthesis of four new voltage-sensitive fluorophores (VoltageFluors, or VFs). Three of these dyes have higher relative voltage sensitivities than the previously-reported indicator, VF2.1.Cl. We pair these new indicators with existing VFs to construct a library of voltage indicators with varying degrees of conjugation between the aniline nitrogen lone pair and the aromatic π system. Using a combination of steady-state and time-resolved fluorescence spectroscopy, cellular electrophysiology, fluorescence lifetime imaging microscopy (FLIM), and functional imaging in mammalian neurons and human cardiomyocytes, we establish a detailed link between the photophysical properties of VF dyes and their ability to report on membrane potential dynamics with high signal-to-noise. Anilines with intermediate degrees of conjugation to the aromatic π system experience intermediate rates of PeT and possess the highest absolute voltage sensitivities. Measured using FLIM in patch-clamped HEK cells, we find that the absolute voltage sensitivity of fluorescence lifetime (∆τfl per mV) provides the best predictor of dye performance in cellular systems.


New Chemical Tools for Exploring Cellular Physiology

National Institute of General Medical Sciences

Find out more...

University of California Berkeley Chemistry-Biology Interface Training Program

National Institute of General Medical Sciences

Find out more...


Email Address of Submitting Author


University of California, Berkeley


United States of America

ORCID For Submitting Author


Declaration of Conflict of Interest

no conflict of interest