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001-es BibID:	BIBFORM077483
Első szerző:	Ujlaky-Nagy László (biofizikus)
Cím:	Flow cytometric FRET analysis of protein interactions / László Ujlaky-Nagy, Péter Nagy, János Szöllősi, György Vereb
Dátum:	2018
Megjegyzések:	In the past decades, investigation of protein?protein interactions in situ in living or intact cells has gained expanding importance as structure/function relationships proposed from bulk biochemistry and molecular modeling experiments required con?rmation at the cellular level. Fo ? rster (?uorescence) resonance energy transfer (FRET)-based methods are excellent tools for determining proximity and supramolecular organization of biomolecules at the cell surface or inside the cell. This could well be the basis for the increasing popularity of FRET. In fact, the number of publications exploiting FRET has exploded since the turn of the millennium. Interestingly, most applications are microscope-based, and only a fraction employs ?ow cytometry, even though the latter offers great statistical power owed to the potentially huge number of individually measured cells. However, with the increased availability of multi-laser ?ow cytometers, strategies to obtain absolute FRET ef?ciencies can now be relatively facilely implemented. In this chapter, we intend to provide generally useable protocols for measuring FRET in ?ow cytometry. After a concise theoretical introduction, recipes are provided for successful labeling techniques and measurement approaches. The simple, quenching-based population-level measurement, the classic ratiometric, intensity-based technique providing cell-by-cell actual FRET ef?ciencies, and a more advanced version of the latter, allowing for cell-by-cell auto?uorescence correction are described. An Excel macro pre-loaded with spectral data of the most commonly used ?uorophores is also provided for easy calculation of average FRET ef?ciencies. Finally, points of caution are given to help design proper experiments and critically interpret the results.
Tárgyszavak:	Orvostudományok Elméleti orvostudományok könyvfejezet Förster resonance energy transfer Fluorescence resonance energy transfer Flow cytometry Protein interactions Molecular proximity FCET
Megjelenés:	Flow Cytometry Protocols / eds. Teresa S. Hawley, Robert G. Hawley. - p. 393-419. -
További szerzők:	Nagy Péter (1971-) (biofizikus) Szöllősi János (1953-) (biofizikus) Vereb György (1965-) (biofizikus, orvos)
Internet cím:	Szerző által megadott URL DOI Intézményi repozitóriumban (DEA) tárolt változat
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