Towards in vivo imaging of intramolecular fluorescence resonance energy transfer parameters

Abstract

Fluorescence resonance energy transfer (FRET) is a nonradiative energy transfer process based on dipole-dipole interaction between donor and acceptor fluorophores that are spatially separated by a distance of a few nanometers. FRET has proved to be of immense value in the study of cellular function and the underlying cause of disease due to, for example, protein misfolding (of consequence in Alzheimer's disease). The standard parameterization in intramolecular FRET is the lifetime and yield, which can be related to the donor-acceptor (DA) distance. FRET imaging has thus far been limited to in vitro or near-surface microscopy because of the deleterious effects of substantial scatter. We show that it is possible to extract the microscopic FRET parameters in a highly scattering environment by incorporating the FRET kinetics of an ensemble of DA molecules connected by a flexible or rigid linker into an optical diffusion tomography (ODT) framework. We demonstrate the efficacy of our approach for extracting the microscopic DA distance through simulations and an experiment using a phantom with scattering properties similar to tissue. Our method will allow the in vivo imaging of FRET parameters in deep tissue, and hence provide a new vehicle for the fundamental study of disease.

In vivo mouse fluorescence imaging for folate-targeted delivery and release kinetics

“…We used a folate-DA conjugate, a surrogate for the folate-drug conjugate, as previous studies have shown that attaching either fluorophores or anti-cancer drugs to folic acid does not interfere with the uptake of folate into cancer cells. To image in deep tissue where tumors occur, we used the diffusion equation to model photon transport in the heavily scattering regime [33,36,37]. This method provides the foundation for a unique and practical tool that can be used to monitor and evaluate pharmacokinetics in preclinical studies.…”

“…FRET [27, 38] is an energy transfer process between donor and acceptor molecules spatially separated by a distance less than 10 nm that can provide invaluable information for nanomedicine. With FODT, the FRET information is accessible in deep tissue [36, 37, 39]. The FRET distance (DA distance) offers a quantitative measure of molecular activity as the donor fluorescence intensity and lifetime are dependent on the FRET distance.…”

Fabrication and application of heterogeneous printed mouse phantoms for whole animal optical imaging

“…In optical diffusion imaging methods such as DOT, polystyrene beads [21], TiO 2 scatterers in acrylic [22], and Intralipid [23,24] have commonly been used as phantoms. However, phantoms created from these materials typically have simple shapes, such as those that can be formed by molds [10].…”

Förster resonance energy transfer photoacoustic microscopy

“…Compared with fluorescence techniques, 5,6 PA imaging of FRET has inherent merits. By utilizing low ultrasonic scattering, PA imaging enables high-resolution, deeply penetrating imaging in biological tissue.…”

“…It can be designed to provide either submicron resolution at a shallow depth or a centimeter penetration depth while a high depth to resolution ratio is maintained. [4][5][6][7][8][9][10] Therefore the ultimate applications of FRET in live cells up to organs in vivo can potentially benefit from the multiscale PA imaging capability. In this work, we demonstrate the initial experiments with PA imaging of FRET in solutions using a dual-modality PA and fluorescence confocal microscope, diagrammed in [ Fig.…”