KEYWORDS: Fluorescence imaging, Modulation, In vivo imaging, Image resolution, Fluorescence, Contrast agents, Tumors, Near infrared, Quantum efficiency, Cancer detection
In vivo tumor imaging and delineating tumor margins serves a critical role in early detection and treatment of cancer. Together with our industrial collaborators, InnoSense LLC (Torrance, CA), we previously have developed ThermoDots™, temperature-responsive micelles that encapsulate an FDA approved near infrared (NIR) imaging agent, indocyanine green (ICG). ThermoDots enhances the signal and enable high resolution fluorescence imaging capability. In this work, they are conjugated with PSMA antibodies for prostate cancer imaging.
We previously introduced a novel method called temperature modulated fluorescence tomography (TMFT), which utilizes temperature sensitive ICG loaded pluronic nanocapsules termed ThermoDots and high intensity focused ultrasound (HIFU). TMFT leverages the superior sensitivity of the fluorescence imaging and the high spatial-resolution of focused ultrasound. Previously, we have presented a prototype system using a CCD camera. However, the scan time was too long for in vivo imaging since HIFU was scanned in a step-and-shoot mode. Here, we present the new continuous line scanning scheme, which drastically reduces the imaging time and paves the wave for in vivo preclinical imaging.
High scattering in biological tissues severely degrades the spatial resolution of optical fluorescence imaging in thick tissue. As one of the most sensitive in vivo molecular imaging modalities, Fluorescence Tomography plays an essential role in preclinical studies. To overcome the limitations of FT, we introduced a novel method termed, temperature modulated fluorescence tomography (TMFT). TMFT is based on two key elements: 1) temperature sensitive fluorescent agent (ThermoDots) and 2) high intensity focused ultrasound (HIFU). TMFT localizes the position of the fluorescent ThermoDots by scanning a HIFU beam across the tissue while monitoring the variation in the measured fluorescence signals. Actually, a binary mask is built by monitoring the sudden jumps in the fluorescence signal corresponding to the HIFU scan over a position containing ThermoDots. This binary map is used as functional a priori during the FT image reconstruction process. TMFT not only allowed us to resolve ThermoDots with high spatial resolution (~1.3 mm), deep in tissue (~ 60 mm) but with high quantitative accuracy as well (< 3% error). In this paper, we present the latest prototype of TMFT. Here, the fluorescence signals are acquired using a CCD camera, which increases the sensitivity of the system compared to the previous fiber-based system.
The strong scattering and absorption of light in biological tissue makes it challenging to model the propagation of light, especially in deep tissue. This is especially true in fluorescent tomography, which aims to recover the internal fluorescence source distribution from the measured light intensities on the surface of the tissue. The inherently ill-posed and underdetermined nature of the inverse problem along with strong tissue scattering makes Fluorescence Tomography (FT) extremely challenging. Previously, multispectral detection fluorescent tomography (FT) has been shown to improve the image quality of FT by incorporating the spectral filtering of biological tissue to provide depth information to overcome the inherent absorption and scattering limitations. We investigate whether multi-wavelength fluorescent tomography can be used to distinguish the signals from multiple fluorophores with overlapping fluorescence spectrums using a unique near-infrared (NIR) swept laser. In this work, a small feasibility study was performed to see whether multi-wavelength FT can be used to detect subtle shifts in the absorption spectrum due to differences in fluorophore microenvironment.
Optical imaging has long been hindered by the high absorption and scattering of light in biological tissue. This makes it difficult to probe beyond a few millimeters beneath the surface without sacrificing image resolution and quantitative accuracy. Strong scattering and the inherent nature of the inverse problem makes fluorescence diffuse optical tomography (FT) extremely challenging. To this end, multi-modality techniques that combine anatomical imaging with the functional optical information have been used to improve the resolution and accuracy of FT. Previously, we have reported on the feasibility of a new imaging method, "Thermal Outlining using Focused Ultrasound" (TOFU), which combines the sensitivity of FT with the resolution of focused ultrasound using temperature reversible fluorescent probes. In this method, the position of the temperature reversible fluorescent probes is localized by an increase in fluorescent signal when the hot spot of the focused ultrasound beam is scanned over the medium. This a priori information is then utilized to guide and constrain conventional reconstruction algorithm to recover the position and concentration of the probes more accurately. The small size of the focal spot (~1.4 mm) up to a depth of 6 cm, allows imaging the distribution of these temperature sensitive agents with not only high spatial resolution but also high quantitative accuracy in deep tissue. In this work, the performance of the system will be evaluated using simulation and phantoms to investigate the dependence that size of the fluorescent distribution has on the TOFU system performance.
Fluorescence tomography is a non invasive, non ionizing imaging technique able to provide a 3D distribution of fluorescent
agents within thick highly scattering mediums, using low cost instrumentation. However, its low spatial resolution due to
undetermined and ill-posed nature of its inverse problem has delayed its integration into the clinical settings. In addition,
the quality of the fluorescence tomography images is degraded due to the excitation light leakage contaminating the
fluorescence measurements. This excitation light leakage results from the excitation photons that cannot be blocked by the
fluorescence filters. In this contribution, we present a new method to remove this excitation light leakage noise based on
the use of a temperature sensitive fluorescence agents. By performing different sets of measurements using this temperature
sensitive agents at multiple temperatures, the excitation light leakage can be estimated and then removed from the
measured fluorescence signals . The results obtained using this technique demonstrate its potential for use in in-vivo small
animal imaging.
Fluorescent tomography has been hindered by poor tissue penetration and weak signal which results in poor spatial resolution and quantification accuracy. Recently, it has been reported that activatable temperature responsive fluorescent probes which respond to focused ultrasound heating can improve the resolution and quantification of fluorescent tomography in deep tissue. This has lead to a new imaging modality, "Temperature-modulated fluorescent tomography." This technique relies on activatable thermo-sensitive fluorescent nanocapsules for whose fluorescence quantum efficiency is temperature dependent. Within a 4-5° C temperature range, the fluorescent signal increase more than 10-fold. In this molecular probe, Indocyanine Green (ICG) is encapsulated inside the core of a thermo-reversible pluronic micelle. Here we show the fluorescence response and temperature range of the nanocapsules which have been optimized for a higher temperature range to be used for in vivo animal imaging. We report on the feasibility of these temperature-sensitive reversible nanocapsules for in vivo applications by studying the pharmacokinetics in a subcutaneous mouse tumor model in vivo.
To overcome the strong scattering in biological tissue that has long afflicted fluorescence tomography, we have
developed a novel technique, "temperature-modulated fluorescence tomography" (TM-FT) to combine the sensitivity of
fluorescence imaging with focused ultrasound resolution. TM-FT relies on two key elements: temperature sensitive ICG
loaded pluronic nanocapsules we termed ThermoDots and high intensity focused ultrasound (HIFU). TM-FT localizes
the position of the fluorescent ThermoDots by irradiating and scanning a HIFU beam across the tissue while
conventional fluorescence tomography measurements are acquired. The HIFU beam produces a local hot spot, in which
the temperature suddenly increases changing the quantum efficiency of the ThermoDots. The small size of the focal spot
(~1 mm) up to a depth of 6 cm, allows imaging the distribution of these temperature sensitive agents with not only high
spatial resolution but also high quantitative accuracy in deep tissue using a proper image reconstruction algorithm.
Previously we have demonstrated this technique with a phantom study with ThermoDots sensitive in the 20-25°C range.
We recently optimized the ThermoDots for physiological temperatures. In this work, we will demonstrate a new HIFU
scanning method which is optimized for in vivo studies. The performance of the system is tested using a phantom that
resembles a small animal bearing a small tumor targeted by ThermoDots.
KEYWORDS: Luminescence, Fluorescence tomography, Ultrasonography, Modulation, Tissues, In vivo imaging, Image resolution, Signal detection, Temperature metrology, Transducers
Low spatial resolution due to strong tissue scattering is one of the main barriers that prevent the wide-spread use of fluorescence tomography. To overcome this limitation, we previously demonstrated a new technique, temperature modulated fluorescence tomography (TM-FT), which relies on key elements: temperature sensitive ICG loaded pluronic nanocapsules and high intensity focused ultrasound (HIFU), to combine the sensitivity of fluorescence imaging with focused ultrasound resolution. While conventional fluorescence tomography measurements are acquired, the tissue is scanned by a HIFU beam and irradiated to produce a local hot spot, in which the temperature increases nearly 5K. The fluorescence emission signal measured by the optical detectors varies drastically when the hot spot overlays onto the location of the temperature dependent nanocapsules. The small size of the focal spot (~1.4 mm) up to a depth of 6 cm, allows imaging the distribution of these temperature sensitive agents with not only high spatial resolution but also high quantitative accuracy in deep tissue using a proper image reconstruction algorithm. Previously we have demonstrated this technique with a phantom study with nanocapsules sensitive to 20-25°C range. In this work, we will show the first nanocapsules optimized for in vivo animal imaging.
It is challenging to image fluorescence objects with high spatial resolution in a highly scattering medium. Recently reported temperature-sensitive indocyanine green-loaded pluronic nanocapsules can potentially alleviate this problem. Here we demonstrate a frequency-domain temperature-modulated fluorescence tomography system that could acquire images at high intensity-focused ultrasound resolution with use of these nanocapsules. The system is experimentally verified with a phantom study, where a 3-mm fluorescence object embedded 2 cm deep in a turbid medium is successfully recovered based on both intensity and lifetime contrast.
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