Journal
OPTICA
Volume 8, Issue 6, Pages 885-897Publisher
Optica Publishing Group
DOI: 10.1364/OPTICA.426870
Keywords
-
Categories
Funding
- National Institutes of Health [R01 CA194697, R01 CA222405, R01 NS107553]
- National Science Foundation [CBET-1554516]
- University of Notre Dame
- Elizabeth and Michael Gallagher Family
- Alfred P. Sloan Foundation
- Berry Family Foundation Graduate Fellowship of Advanced Diagnostics Therapeutics
Ask authors/readers for more resources
Instant FLIM is a method of analog signal processing that allows real-time streaming of fluorescence intensity, lifetime, and phasor imaging data, improving the efficiency and real-time capabilities of FLIM techniques. By upgrading existing two-photon microscopes with cost-effective components and open-source software, this technology can be easily implemented, along with enhancements in functionality through phasor segmentation, adaptive optics, and super-resolution techniques.
Traditional fluorescence microscopy is blind to molecular microenvironment information that is present in a fluorescence lifetime, which can be measured by fluorescence lifetime imaging microscopy (FLIM). However, most existing FLIM techniques are slow to acquire and process lifetime images, difficult to implement, and expensive. Here we present instant FLIM, an analog signal processing method that allows real-time streaming of fluorescence intensity, lifetime, and phasor imaging data through simultaneous image acquisition and instantaneous data processing. Instant FLIM can be easily implemented by upgrading an existing two-photon microscope using cost-effective components and our open-source software. We further improve the functionality, penetration depth, and resolution of instant FLIM using phasor segmentation, adaptive optics, and super-resolution techniques. We demonstrate through-skull intravital 3D FLIM of mouse brains to depths of 300 mu m and present the first in vivo 4D FLIM of microglial dynamics in intact and injured zebrafish and mouse brains for up to 12 h. (C) 2021 Optical Society of America under the terms of the OSA Open Access Publishing Agreement
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available