Abstract
The effects of donor homo-Förster resonance energy transfer (homo-FRET) taking place in hetero-FRET systems is described in the context of hetero-FRET detection via donor and acceptor fluorescence anisotropies in cell surface receptor clusters. Donor homo-FRET can influence both the efficiency of detection as well as the magnitude of the detectable hetero-FRET. A 4-fold polarized FRET detection scheme-tetrapolarization FRET (4polFRET)-is proposed not only for discriminating the effects of homo-FRET from those of hetero-FRET, but also for correlating homo-associations of the donors and acceptors at different donor-acceptor distances, even beyond the critical Förster distance for hetero-FRET ( R0). The method is based on suppressing homo-FRET at the donor side with red-edge excitation. After the anisotropy effects of physical rotation and homo-FRET were separated by site-selective spectroscopy, the magnitude of the effect of homo-FRET on hetero-FRET has been estimated. It has been found significant, offering a new sensitive technique for detecting conformational dynamics via the homo-FRET mediated component of hetero-FRET, the "homo-FRET enhanced hetero-FRET" or "homo-FRET gate". The method is realizable in flow, as well as in image cytometry equipped with polarization detecting facility.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.