Two-photon FLIM-FRET microscopy for protein localization

Ye Chen; Ammasi Periasamy

doi:10.1117/12.538312

21 June 2004 Two-photon FLIM-FRET microscopy for protein localization

Ye Chen, Ammasi Periasamy

Proceedings Volume 5323, Multiphoton Microscopy in the Biomedical Sciences IV; (2004) https://doi.org/10.1117/12.538312
Event: Biomedical Optics 2004, 2004, San Jose, CA, United States

Abstract

Intensity based FRET visualizes protein molecules in living cells and tissues. Lifetime techniques, however, will demonstrate the dynamic functional activity of the protein molecules, because its signal does not depend on changes in fluorophore concentration or excitation intensity. If a laser pulse excites a large number of similar molecules with a similar local environment, and as long as no energy is transferred to another molecule, the lifetime is the “natural fluorescence lifetime”. If energy is transferred, however, the actual fluorescence lifetime is less than the natural lifetime, because an additional path for de-excitation is present. With the occurrence of FRET, strong energy transfer results in extreme quenching of the donor fluorescence and a decrease in the fluorescence lifetime. In this paper we will explain the development of the two-photon FLIM-FRET microscopy with our existing multiphoton microscopy and demonstrate the change in donor lifetime by photobleaching the acceptor molecules in living cells.

Citation Download Citation

Ye Chen and Ammasi Periasamy "Two-photon FLIM-FRET microscopy for protein localization", Proc. SPIE 5323, Multiphoton Microscopy in the Biomedical Sciences IV, (21 June 2004); https://doi.org/10.1117/12.538312

ACCESS THE FULL ARTICLE

INSTITUTIONAL
Select your institution to access the SPIE Digital Library.

SELECT YOUR INSTITUTION

PERSONAL
Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.

PERSONAL SIGN IN

No SPIE Account? Create one

PURCHASE THIS CONTENT

SUBSCRIBE TO DIGITAL LIBRARY

50 downloads per 1-year subscription

Members: $195

Non-members: $335 ADD TO CART

25 downloads per 1 - year subscription

Members: $145

Non-members: $250 ADD TO CART

PURCHASE SINGLE ARTICLE

Includes PDF, HTML & Video, when available