Fluorescence-lifetime imaging microscopy

"FLIM" redirects here. For other uses, see Flim.

Fluorescence-lifetime imaging microscopy or FLIM is an imaging technique based on the differences in the exponential decay rate of the photon emission of a fluorophore from a sample. It can be used as an imaging technique in confocal microscopy, two-photon excitation microscopy, and multiphoton tomography.

The fluorescence lifetime (FLT) of the fluorophore, rather than its intensity, is used to create the image in FLIM. Fluorescence lifetime depends on the local micro-environment of the fluorophore, thus precluding any erroneous measurements in fluorescence intensity due to change in brightness of the light source, background light intensity or limited photo-bleaching. This technique also has the advantage of minimizing the effect of photon scattering in thick layers of sample. Being dependent on the micro-environment, lifetime measurements have been used as an indicator for pH,[1] viscosity[2] and chemical species concentration.[3][4]

  1. ^ Nakabayashi, Takakazu; Wang, Hui-Ping; Kinjo, Masataka; Ohta, Nobuhiro (4 June 2008). "Application of fluorescence lifetime imaging of enhanced green fluorescent protein to intracellular pH measurements". Photochemical & Photobiological Sciences. 7 (6): 668–670. doi:10.1039/B800391B. ISSN 1474-9092. PMID 18528549. S2CID 42881416.
  2. ^ Levitt, James A.; Kuimova, Marina K.; Yahioglu, Gokhan; Chung, Pei-Hua; Suhling, Klaus; Phillips, David (9 July 2009). "Membrane-Bound Molecular Rotors Measure Viscosity in Live Cells via Fluorescence Lifetime Imaging". The Journal of Physical Chemistry C. 113 (27): 11634–11642. doi:10.1021/jp9013493. hdl:10044/1/15590. ISSN 1932-7447. S2CID 96097931.
  3. ^ Ruedas-Rama, Maria J.; Orte, Angel; Hall, Elizabeth A. H.; Alvarez-Pez, Jose M.; Talavera, Eva M. (20 February 2012). "A chloride ion nanosensor for time-resolved fluorimetry and fluorescence lifetime imaging". Analyst. 137 (6): 1500–1508. Bibcode:2012Ana...137.1500R. doi:10.1039/C2AN15851E. ISSN 1364-5528. PMID 22324050.[permanent dead link]
  4. ^ Agronskaia, Alexandra V.; Tertoolen, L.; Gerritsen, Hans C. (November 2004). "Fast fluorescence lifetime imaging of calcium in living cells". Journal of Biomedical Optics. 9 (6): 1230–1237. Bibcode:2004JBO.....9.1230A. doi:10.1117/1.1806472. ISSN 1083-3668. PMID 15568944.