Fluorescence Lifetime Imaging of Apoptosis

Tomography : a Journal for Imaging Research
Annie XiaoGary D Luker

Abstract

Genetically-encoded fluorescence resonance energy transfer (FRET) reporters are powerful tools to analyze cell signaling and function at single cell resolution in standard two-dimensional cell cultures, but these reporters rarely have been applied to three-dimensional environments. FRET interactions between donor and acceptor molecules typically are determined by changes in relative fluorescence intensities, but wavelength-dependent differences in absorption of light complicate this analysis method in three-dimensional settings. Here we report fluorescence lifetime imaging microscopy (FLIM) with phasor analysis, a method that displays fluorescence lifetimes on a pixel-wise basis in real time, to quantify apoptosis in breast cancer cells stably expressing a genetically encoded FRET reporter. This microscopic imaging technology allowed us to identify treatment-induced apoptosis in single breast cancer cells in environments ranging from two-dimensional cell culture, spheroids with cancer and bone marrow stromal cells, and living mice with orthotopic human breast cancer xenografts. Using this imaging strategy, we showed that combined metabolic therapy targeting glycolysis and glutamine pathways significantly reduced overall breast ...Continue Reading

Citations

Sep 14, 2016·Seminars in Cell & Developmental Biology·Mengjia QianYunfeng Cheng
Jul 1, 2017·Methods and Applications in Fluorescence·Franz-Josef Meyer-Almes
Mar 4, 2018·Journal of Biomedical Optics·Victoria ZherdevaAlexander P Savitsky
Dec 12, 2017·Current Protocols in Cell Biology·Johanna M BuschhausGary D Luker
Oct 13, 2018·Lasers in Medical Science·Aparna Dravid U, Nirmal Mazumder
Jan 21, 2020·Biotechnology and Bioengineering·A H Rezwanuddin AhmedSihong Wang
Aug 5, 2017·Molecular Cancer Therapeutics·Stefanie GalbánBrian D Ross
Jun 18, 2021·Nano-micro Letters·Yuzhen OuyangMinghua Wu

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Methods Mentioned

BETA
imaging techniques
fluorescence resonance
FRET
xenografts
PCR
flow cytometry
transfection
Fluorescence
fluorescence microscopy
fluorescence imaging

Software Mentioned

GraphPad Prism

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