Assessment of Gate Width Size on Lifetime-Based Förster Resonance Energy Transfer Parameter Estimation

Photonics

Sez-Jade Chen¹  Nattawut Sinsuebphon¹ 

[1] Department of Biomedical Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA;

关键词: Förster Resonance Energy Transfer (FRET);    time-resolved imaging;    gate width;    fluorescence lifetime;    gated ICCD;    near infrared (NIR) dyes;    in vivo imaging;   

DOI  :  10.3390/photonics2041027

来源: mdpi

【 摘 要 】

Förster Resonance Energy Transfer (FRET) enables the observation of interactions at the nanoscale level through the use of fluorescence optical imaging techniques. In FRET, fluorescence lifetime imaging can be used to quantify the fluorescence lifetime changes of the donor molecule, which are associated with proximity between acceptor and donor molecules. Among the FRET parameters derived from fluorescence lifetime imaging, the percentage of donor that interacts with the acceptor (in proximity) can be estimated via model-based fitting. However, estimation of the lifetime parameters can be affected by the acquisition parameters such as the temporal characteristics of the imaging system. Herein, we investigate the effect of various gate widths on the accuracy of estimation of FRET parameters with focus on the near-infrared spectral window. Experiments were performed in silico, in vitro, and in vivo with gate width sizes ranging from 300 ps to 1000 ps in intervals of 100 ps. For all cases, the FRET parameters were retrieved accurately and the imaging acquisition time was decreased three-fold. These results indicate that increasing the gate width up to 1000 ps still allows for accurate quantification of FRET interactions even in the case of short lifetimes such as those encountered with near-infrared FRET pairs.

【 授权许可】

CC BY   
© 2015 by the authors; licensee MDPI, Basel, Switzerland.