ArticlePloS one2018
Accepting from the best donor; analysis of long-lifetime donor fluorescent protein pairings to optimise dynamic FLIM-based FRET experiments.
Article in PloS one, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 28 papers.
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Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
28 citing papers in PubMed.
- Tuning the Response of GPCR-Based Yeast Sensors Using Fluorescent Reporters.ACS synthetic biology · 2026Article
- Linking molecular tension and cellular tractions: a multiscale approach to focal adhesion mechanics.Communications biology · 2026Article
- Unravelling molecular dynamics in living cells: Fluorescent protein biosensors for cell biology.Journal of microscopy · 2025Review
- Diverting glial glycolytic flux towards neurons is a memory-relevant role of Drosophila CRH-like signalling.Nature communications · 2024Article
- Article
- Interplay of EXO70 and MLO proteins modulates trichome cell wall composition and susceptibility to powdery mildew.The Plant cell · 2024Article
- Particle-based phasor-FLIM-FRET resolves protein-protein interactions inside single viral particles.Biophysical reports · 2023Article
- Three-Fluorophore FRET Enables the Analysis of Ternary Protein Association in Living Plant Cells.Plants (Basel, Switzerland) · 2022Article
- Stress routes clients to the proteasome via a BAG2 ubiquitin-independent degradation condensate.Nature communications · 2022Article
- Genetic encoding of a highly photostable, long lifetime fluorescent amino acid for imaging in mammalian cells.Chemical science · 2021Article
- FLIM as a Promising Tool for Cancer Diagnosis and Treatment Monitoring.Nano-micro letters · 2021Review
- Single-cell resolved imaging reveals intra-tumor heterogeneity in glycolysis, transitions between metabolic states, and their regulatory mechanisms.Cell reports · 2021Article
- Super-resolution time-resolved imaging using computational sensor fusion.Scientific reports · 2021Article
- A Combined Acceptor Photobleaching and Donor Fluorescence Lifetime Imaging Microscopy Approach to Analyze Multi-Protein Interactions in Living Cells.Frontiers in molecular biosciences · 2021Article
- Fluorescence lifetime imaging with a megapixel SPAD camera and neural network lifetime estimation.Scientific reports · 2020Article
- Cellular Plasticity during Metastasis: New Insights Provided by Intravital Microscopy.Cold Spring Harbor perspectives in medicine · 2020Review
- Mitochondrial pH Nanosensors for Metabolic Profiling of Breast Cancer Cell Lines.International journal of molecular sciences · 2020Article
- How many photons are needed for FRET imaging?Biomedical optics express · 2020Article
- Live-cell FLIM-FRET using a commercially available system.Methods in cell biology · 2020Article
- Comparing the performance of mScarlet-I, mRuby3, and mCherry as FRET acceptors for mNeonGreen.PloS one · 2020Article
Corrections and comments
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Authors and funding
8 authors.
Funding
Abstract
FRET biosensors have proven very useful tools for studying the activation of specific signalling pathways in living cells. Most biosensors designed to date have been predicated on fluorescent protein pairs that were identified by, and for use in, intensity based measurements, however fluorescence lifetime provides a more reliable measurement of FRET. Both the technology and fluorescent proteins available for FRET have moved on dramatically in the last decade. Lifetime imaging systems have become increasingly accessible and user-friendly, and there is an entire field of biology dedicated to refining and adapting different characteristics of existing and novel fluorescent proteins. This growing pool of fluorescent proteins includes the long-lifetime green and cyan fluorescent proteins Clover and mTurquoise2, the red variant mRuby2, and the dark acceptor sREACh. Here, we have tested these donors and acceptors in appropriate combinations against the standard or recommended norms (EGFP and mTFP as donors, mCherry and either Ypet or Venus as acceptors) to determine if they could provide more reliable, reproducible and quantifiable FLIM-FRET data to improve on the dynamic range compared to other donors and breadth of application of biosensor technologies. These tests were performed for comparison on both a wide-field, frequency domain system and a multiphoton, TCSPC time domain FLIM system. Clover proved to be an excellent donor with extended dynamic range in combination with mCherry on both platforms, while mRuby2 showed a high degree of variability and poor FRET efficiencies in all cases. mTFP-Venus was the most consistent cyan-yellow pair between the two FLIM methodologies, but mTurquoise2 has better dynamic range and transfers energy consistently over time to the dark acceptor sRCh. Combination of mTFP-sRCh with Clover-mCherry would allow the simultaneous use of two FLIM-FRET biosensors within one sample by eliminating the crosstalk between the yellow acceptor and green donor emissions.
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