title: Spatial light modulator based active wide-field illumination for ex vivo and in vivo quantitative NIR FRET imaging.
abstract: Fluorescence lifetime imaging is playing an increasing role in drug development by providing a sensitive method to monitor drug delivery and receptor-ligand interactions. However, the wide dynamic range of fluorescence intensity emitted by ex vivo and in vivo samples presents challenges in retrieving information over the whole subject accurately and quantitatively. To overcome this challenge, we developed an active wide-field illumination (AWFI) strategy based on a spatial light modulator that acquires optimal fluorescence signals by enhancing the dynamic range, signal to noise ratio, and estimation of lifetime-based parameters. We demonstrate the ability of AWFI to estimate Förster resonance energy transfer (FRET) donor fraction from dissected organs with high accuracy (standard deviation <6%) over the whole field of view, in contrast with the homogenous wide-field illumination. We further report its successful application to quantitative FRET imaging in a live mouse. AWFI allows improved detection of weak signals and enhanced quantitative accuracy in ex vivo and in vivo molecular fluorescence quantitative imaging. The technique allows for robust quantitative estimation of the bio-distribution of molecular probes and lifetime-based parameters over an extended imaging field exhibiting a large range of fluorescence intensities and at a high acquisition speed (less than 1 min).
authors.affiliation: Department of Biomedical Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA.
authors.forename: Lingling
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Center for Cardiovascular Sciences, Albany Medical College, 43 New Scotland Avenue, Albany, NY 12208, USA.
authors.forename: Ken
authors.initials: K
authors.lastname: Abe
authors.affiliation: Center for Cardiovascular Sciences, Albany Medical College, 43 New Scotland Avenue, Albany, NY 12208, USA.
authors.forename: Shilpi
authors.initials: S
authors.lastname: Rajoria
authors.affiliation: Department of Biomedical Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA.
authors.forename: Qi
authors.initials: Q
authors.lastname: Pian
authors.affiliation: Center for Cardiovascular Sciences, Albany Medical College, 43 New Scotland Avenue, Albany, NY 12208, USA.
authors.forename: Margarida
authors.initials: M
authors.lastname: Barroso
authors.affiliation: Department of Biomedical Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA.
authors.forename: Xavier
authors.initials: X
authors.lastname: Intes
keywords: (170.2945) Illumination design; (170.3650) Lifetime-based sensing; (170.6920) Time-resolved imaging; (230.6120) Spatial light modulators; (260.2160) Energy transfer; (260.2510) Fluorescence