scholarly journals Lightsheet fluorescence lifetime imaging microscopy with wide‐field time‐correlated single photon counting

2019 ◽  
Vol 13 (2) ◽  
Author(s):  
Liisa M. Hirvonen ◽  
Jakub Nedbal ◽  
Norah Almutairi ◽  
Thomas A. Phillips ◽  
Wolfgang Becker ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Single Photon ◽  
Photon Counting ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Wide Field ◽  
Fluorescence Lifetime Imaging ◽  
Single Photon Counting ◽  
Lifetime Imaging

scholarly journals Optical Estimation of Absolute Membrane Potential Using One- and Two-Photon Fluorescence Lifetime Imaging Microscopy

2021 ◽  
Author(s):  
Julia R. Lazzari-Dean ◽  
Evan W. Miller
Keyword(s):  
Membrane Potential ◽  
Fluorescence Lifetime ◽  
Single Photon ◽  
Photon Counting ◽  
Fluorescence Lifetime Imaging ◽  
Single Photon Counting ◽  
Two Photon ◽  
Lifetime Imaging ◽  
Wide Range ◽  
Relative Measurements

AbstractBackgroundMembrane potential (Vmem) exerts physiological influence across a wide range of time and space scales. To study Vmem in these diverse contexts, it is essential to accurately record absolute values of Vmem, rather than solely relative measurements.Materials & MethodsWe use fluorescence lifetime imaging of a small molecule voltage sensitive dye (VF2.1.Cl) to estimate mV values of absolute membrane potential.ResultsWe test the consistency of VF2.1.Cl lifetime measurements performed on different single photon counting instruments and find that they are in striking agreement (differences of <0.5 ps/mV in the slope and <50 ps in the y-intercept). We also demonstrate that VF2.1.Cl lifetime reports absolute Vmem under two-photon (2P) illumination with better than 20 mV of Vmem resolution, a nearly 10-fold improvement over other lifetime-based methods.ConclusionsWe demonstrate that VF-FLIM is a robust and portable metric for Vmem across imaging platforms and under both one-photon and two-photon illumination. This work is a critical foundation for application of VF-FLIM to record absolute membrane potential signals in thick tissue.

Fluorescence lifetime imaging by time-correlated single-photon counting

2003 ◽  
Vol 63 (1) ◽  
pp. 58-66 ◽  
Author(s):  
W. Becker ◽  
A. Bergmann ◽  
M.A. Hink ◽  
K. König ◽  
K. Benndorf ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Single Photon ◽  
Photon Counting ◽  
Fluorescence Lifetime Imaging ◽  
Single Photon Counting ◽  
Lifetime Imaging
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