title: Measuring NLR Oligomerization IV: Using Förster Resonance Energy Transfer (FRET)-Fluorescence Lifetime Imaging Microscopy (FLIM) to Determine the Close Proximity of Inflammasome Components.
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.
authors.affiliation: Institute for Research in Biomedicine (IRB Barcelona), Parc Científic de Barcelona, Baldiri Reixac 10, 08028, Barcelona, Spain.
authors.forename: Catrin
authors.initials: C
authors.lastname: Youssif
authors.affiliation: Memory Unit, Neurology Department, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain.
authors.forename: Bárbara
authors.initials: B
authors.lastname: Flix
authors.affiliation: Memory Unit, Neurology Department, Hospital de la Santa Creu i Sant Pau, Barcelona, Spain.|Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERned), Madrid, Spain.
authors.forename: Olivia
authors.initials: O
authors.lastname: Belbin
authors.affiliation: Institute for Research in Biomedicine (IRB Barcelona), Parc Científic de Barcelona, Baldiri Reixac 10, 08028, Barcelona, Spain. monica.comalada@irbbarcelona.org.
authors.forename: Mònica
authors.initials: M
authors.lastname: Comalada
mesh_terms: D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D058847:Inflammasomes / Q000737:chemistry / Q000378:metabolism*; D008856:Microscopy, Fluorescence / Q000379:methods; D000070576:NLR Proteins / Q000737:chemistry* / Q000378:metabolism; D025941:Protein Interaction Mapping; D055503:Protein Multimerization
chemical_list: D058847:Inflammasomes; D000070576:NLR Proteins
keywords: Caspase-1; Endogenous proteins; FLIM; FRET; Immunofluorescence; Inflammasome; Macrophages; Multi-protein complex; NLRP3