Abstract
We describe a red-shifted fluorescence resonance energy transfer (FRET) pair optimized for dual-color fluorescence lifetime imaging (FLIM). This pair utilizes a newly developed FRET donor, monomeric cyan-excitable red fluorescent protein (mCyRFP1), which has a large Stokes shift and a monoexponential fluorescence lifetime decay. When used together with EGFP-based biosensors, the new pair enables simultaneous imaging of the activities of two signaling molecules in single dendritic spines undergoing structural plasticity.
🔬 Techniques
✨ Fluorophores
🧪 Sample Preparation
🔬 Cell Lines
🏛️ Research Organizations (ROR)
Affiliated research institutions:
📊 Figures
Figure 1
Characterization of CyRFP1 and dual-imaging with GCaMP6
(a) Normalized absorbance spectra of mCyRFP1, CyRFP1, and mEGFP. Dotted line indicates approximate 2p excitation (920 nm). (b) Normalized emission spectra of mCyRFP1, CyRFP1, and mEGFP. Dotted rectang...
Figure 2
Characterization of the mCyRFP1-mMaroon1 FRET pair for FLIM
(a) Normalized (norm.) spectra of absorptions and excitations (ex, excitation; em, emission) of mCyRFP1 and mMaroon1. (b) Fluorescence lifetime decay curves of dominant negative (DN) and constitutive ...
Figure 3
Simultaneous FLIM measurements of RhoA and CaMKII activation in single dendritic spines undergoing structural plasticity
(a) Schematics of CaMKII-alpha sensor (Green-Camuiu03b1) and RhoAu2013CyRM used for experiments. (b,c) Representative fluorescence lifetime images of RhoA (b) and CaMKII (c) activation in a dendritic ...
Figure images are served from the NIH/NLM PubMed Central Open Access Subset or Europe PMC; copyright remains with the publishers and authors.
💬 Discussion
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