Abstract
Intracellular signaling and cellular activation have been demonstrated to reside on multi-protein complexes rather than in isolated proteins. Consequently, techniques to resolve these complexes have gained much attention over the last few years. Förster Resonance Energy Transfer (FRET) coupled with Fluorescence Lifetime Imaging Microscopy (FLIM) is a powerful tool to discriminate direct interactions between two proteins within a multi-protein complex. Here, we present the use of FRET-FLIM as an experimental tool for the interpretation of the inflammasome composition. We also introduce some considerations required for the correct use of this technique and the control experiments that should be implemented.
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Acknowledgement
M.C. and O.B. contributed equally to this work. The authors thank Carmen Casal and Eva Companys for their help in developing the FRET-FLIM technique presented here. O.B is a recipient of the “Miguel Servet” Grant provided by the FEDER (European Funds for Regional Development) and the Carlos III Institute of Health (Ministry for Economy and Competitivity, Spain). C.Y. is recipient of “La Caixa” fellowship.
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Youssif, C., Flix, B., Belbin, O., Comalada, M. (2016). Measuring NLR Oligomerization IV: Using Förster Resonance Energy Transfer (FRET)-Fluorescence Lifetime Imaging Microscopy (FLIM) to Determine the Close Proximity of Inflammasome Components. In: Di Virgilio, F., Pelegrín, P. (eds) NLR Proteins. Methods in Molecular Biology, vol 1417. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3566-6_11
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DOI: https://doi.org/10.1007/978-1-4939-3566-6_11
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