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Event Type:  Seminar (This is an NIH Science event)
Annual Lecture:  Other
Title:  Fluorescence Lifetime imaging microscopy (FLIM) for Quantitative Live-cell Measurements with Forster Resonance Energy Transfer (FRET) Probes
Description:  Most of the commonly used fluorescence microscopy techniques today depend on the detection of the emission intensity of various fluorescent proteins, dyes or endogenous reports. In addition to the crucially useful separation of the emission and excitation wavelengths, the fluorescence process offers another exciting and so far underused opportunity to explore living cells through the fluorescence lifetimes of fluorophores. The fluorescence lifetime, which is independent of the emission intensity, is a measure of the time it takes before the excited fluorophore returns to its ground state. The interesting feature of fluorescence is that the energy dissipation from the excited state depends on the molecular condition of the fluorophores (protonation, oxygenation, etc.) and their environment (such as binding to fluorescent or non-fluorescent molecules). Because of that, the fluorescence lifetime imaging microscopy (FLIM) has the potential to provide real-time measurements of molecular interactions and biochemical reactions in live cells.
The time-correlated single photon counting (TCSPC) method of FLIM provides better lifetime resolution and higher photon usage efficiency than its main alternative, the fluorescence polarization FLIM. We used the TCSPC FLIM with monomolecular FRET reporters to study the role of the small GTPase Ran in the regulation of mitosis, cell cycle or DNA repair. In those studies, the quantitative properties of the FLIM/FRET imaging gave us the particularly useful insights. FLIM provides a rigorous and quantitative method of FRET measurements and has several distinct advantages over the intensity-based methods, including relaxed concerns for spectral crosstalks (only donor emission is detected), insensitivity to sensor concentration (within reasonable limits) and the straightforward relationship between the measured lifetimes and FRET efficiency. The main limitations in FLIM/FRET, which should not be underestimated, is the need for a large num
Series Name:  Light Microscopy Interest Group Seminar
Videocast:  Event will not be videocast
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Tuesday, October 17, 2017   11:00am - 12:00pm Add To Outlook Calendar     Add To iCal Calendar     Add To Entourage Calendar
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Name:   Petr Kalab
Title:   Dr
Organization:   Johns Hopkins U
City/Province:   Baltimore
State:   Maryland
Country:   USA

Organization(s):  [NIH] Light Microscopy Interest Group

Location:  On the main NIH Campus
Building:  37
Room:  4107/4041
Street Address:  37 Convent Drive
City:  Bethesda
State:  Maryland
Zip Code:  20906

Name:   Tatiana Karpova
Phone:   240-760-6637
Fax:   240-541-4450
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