ArticleJournal of biomedical optics2012
Methodological considerations for global analysis of cellular FLIM/FRET measurements.
Article in Journal of biomedical optics, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 6 papers.
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Who cites it
6 citing papers in PubMed.
- Development and Characterization of a FRET-based Formin Tension Sensor in Living Cells.bioRxiv : the preprint server for biology · 2026Article
- Blind Resolution of Lifetime Components in Individual Pixels of Fluorescence Lifetime Images Using the Phasor Approach.The journal of physical chemistry. B · 2020Article
- Single cell-based fluorescence lifetime imaging of intracellular oxygenation and metabolism.Redox biology · 2020Article
- Quantifying intracellular protein binding thermodynamics during mechanotransduction based on FRET spectroscopy.Methods (San Diego, Calif.) · 2014Article
- τFCS: multi-method global analysis enhances resolution and sensitivity in fluorescence fluctuation measurements.PloS one · 2014Article
- Quantitative tomographic imaging of intermolecular FRET in small animals.Biomedical optics express · 2012Article
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Authors and funding
4 authors.
Funding
Abstract
Global algorithms can improve the analysis of fluorescence energy transfer (FRET) measurement based on fluorescence lifetime microscopy. However, global analysis of FRET data is also susceptible to experimental artifacts. This work examines several common artifacts and suggests remedial experimental protocols. Specifically, we examined the accuracy of different methods for instrument response extraction and propose an adaptive method based on the mean lifetime of fluorescent proteins. We further examined the effects of image segmentation and a priori constraints on the accuracy of lifetime extraction. Methods to test the applicability of global analysis on cellular data are proposed and demonstrated. The accuracy of global fitting degrades with lower photon count. By systematically tracking the effect of the minimum photon count on lifetime and FRET prefactors when carrying out global analysis, we demonstrate a correction procedure to recover the correct FRET parameters, allowing us to obtain protein interaction information even in dim cellular regions with photon counts as low as 100 per decay curve.
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