Design and characterization of a time-domain optical tomography platform for mesoscopic lifetime imaging

Abstract

We report on the system design and instrumental characteristics of a novel time-domain mesoscopic fluorescence molecular tomography (TD-MFMT) system for multiplexed molecular imaging in turbid media. The system is equipped with a supercontinuum pulsed laser for broad spectral excitation, based on a high-density descanned raster scanning intensity-based acquisition for 2D and 3D imaging and augmented with a high-dynamical range linear time-resolved single-photon avalanche diode (SPAD) array for lifetime quantification. We report on the system’s spatio-temporal and spectral characteristics and its sensitivity and specificity in controlled experimental settings. Also, a phantom study is undertaken to test the performance of the system to image deeply-seated fluorescence inclusions in tissue-like media. In addition, ex vivo tumor xenograft imaging is performed to validate the system’s applicability to the biological sample. The characterization results manifest the capability to sense small fluorescence concentrations (on the order of nanomolar) while quantifying fluorescence lifetimes and lifetime-based parameters at high resolution. The phantom results demonstrate the system’s potential to perform 3D multiplexed imaging thanks to spectral and lifetime contrast in the mesoscopic range (at millimeters depth). The ex vivo imaging exhibits the prospect of TD-MFMT to resolve intra-tumoral heterogeneity in a depth-dependent manner.

Characterization of fluorescence lifetime of organic fluorophores for molecular imaging in the SWIR window

“…For the SWIR-specific fluorophores, we employed commercially available probes, IR-E1050 and IR-T1050 (Nirmidas Biotech). 2 Compared to the diagram reported previously, 11 TD-MFMT system was adapted by replacing the sCMOS with the InGaAs camera (Owl 640 II, Raptor Photonics) and switching the optics to the SWIR-optimized components (shown in Fig. 1) for optimal detection efficiency.…”

“…17, 18 One can follow the workflow of the TD data collection and lifetime quantification laid out in our previous work. 11 WF and SPAD-based intensities measured by the InGaAs camera and the SPAD, the lifetime quantification results, as well as the normalized IRF and representative decays of four fluorophores are shown in Fig. 2 A and C.…”

“…9,10 Integrating FLI into well-established intensity-based imaging systems can complement their performances; recently, our group characterized and validated scanning-based Time-domain Mesoscopic Fluorescence Molecular Tomography (TD-MFMT), capable of acquiring fluorescence intensity for 3D probe reconstruction as well as resolving fluorescence lifetime information in the visible-NIR window. 11 Nevertheless, challenges in implementing FLI in SWIR arise from limitations in both SWIR-specific probes and detectors. Current studies mostly use inorganic probes that can be used for luminescence lifetime multiplexing 12 but which raise biocompatibility issues.…”

Ex vivo mesoscopic assessment of drug delivery and target engagement via lifetime FRET

“…2 As a novel fluorescence optical imaging technique, time-domain mesoscopic fluorescence molecular tomography (TD-MFMT) has been recently proposed to report on fluorescence expression in 3D and target engagement via lifetime-based Förster resonance energy transfer (FRET) in biotissues at mesoscopic regime (millimeters depth) and with higher resolution (hundreds of microns). 3 Herein, we report on preliminary results acquired by TD-MFMT for ex vivo mesoscopic assessing drug delivery and target engagement for HER2-positive breast tumor (AU565) and ovarian tumor (SK-OV-3) xenograft models, which were extracted from nude mice injected with AF750-trastuzumab and AF700-Trastuzumab conjugates (NIR FRET pair).…”

Characterization of fluorescence lifetime of organic fluorophores for molecular imaging in the shortwave infrared window

“…9,10 Integrating FLI into well-established intensity-based imaging systems can complement their performances; recently, our group characterized and validated scanning-based time-domain mesoscopic fluorescence molecular tomography (TD-MFMT), capable of acquiring fluorescence intensity for 3D probe reconstruction as well as resolving fluorescence lifetime information in the visible-NIR window. 11 Nevertheless, challenges in implementing FLI in SWIR arise from limitations in both SWIR-specific probes and detectors. Current studies mostly use inorganic probes that can be used for luminescence lifetime multiplexing 12 but that raise biocompatibility issues.…”